Clinical Chemistry Notes With Blanks

CLINICAL CHEMISTRY MTAP
BY: GEORGE VINCENT GELLENA, RMT

Remedios Trinidad Romualdez Medical Foundation
CONTENTS: Table 1. – Types of Error

1 Quality Management Random Error Systematic Error
2 Automation Description
4 Carbohydrates
7 Lipids
10 Proteins
12 Enzymes
16 Non-Protein Nitrogens
18 Liver Function Tests Examples
20 Electrolytes
23 Acid-Base Balance
24 Endocrinology
30 Therapeutic Drug Monitoring
32 Toxicology
QUALITY MANAGEMENT
 ___________________ - Is a system of ensuring accuracy
Statistics
& precision in the laboratory by including quality control
 ____________________ – measure of central tendency;
reagents in every series of measurements
measure of accuracy; AVERAGE
 ___________________- is a systematic action necessary
 ____________________ – measure of dispersion of
to provide adequate confidence that laboratory services
values from the mean; measure of precision; most
will satisfy the given medical needs for patient care
frequently used measure of variation
 ___________________ – material of known
 ____________________ - index of precision; percentile
concentration used in developing a standard curve
expression of the mean
and/or instrument calibration
 ____________________ - measure of variability
 ___________________ – sample of known quantity with
 ____________________ - determines whether there is a
several analytes present
statistically significant difference between the standard
Parameters
deviations of two groups of data
 ___________________ - Is the Ability of an analytical
 ____________________ - determines whether there is a
method to measure the smallest concentration of the
statistically significant difference between the means of
analyte of interest
two groups of data
 ___________________ - Is the Ability of an analytical
 ____________________ - MIDPOINT of the distribution;
method to measure ONLY the analyte of Interest
value of the observation that divides the observation into
 ___________________ - Is the Nearness or Closeness of
two equal groups of data
the Assayed value to the true or target value
 ____________________ – most FREQUENT observation
 ___________________- The ability of an analytical
 ____________________ - is the difference between the
method to give repeated results on the same sample that
highest and lowest score in data
agree with one another
 ___________________ - The degree by w/c a method is Quality Control Charts
 ____________________ - data element are centered
easily repeated.
 ___________________ - The ability of an analytical around the mean with most elements close to the mean
method to maintain accuracy & precision over an  ____________________ - provides the earliest
extended period of time during w/c equipment, indication of systematic error (trend); requires computer
reagents, & personnel may change implementation
 ___________________ - The Ability of an analytical  ____________________ – compare results obtained on
method to detect the proportion of individuals with the a high and low control serum from different laboratories
disease. (Screening tests require high sensitivity)  ___________________________ – most widely used QC
 ___________________ - The Ability of an analytical chart in the clinical laboratory; allows laboratorians to
method to detect the proportion of individuals without apply multiple rules without the aid of computer;
the disease. (Confirmatory tests require high specificity) identifies both random and systematic error
 ___________________ – highest frequency occurs with Table 2. Errors observed in LJ Chart
the use of handwritten labels and request forms TREND SHIFT
COLLEGE OF MEDICAL LABORATORY SCIENCE | 1 | 34

Clinical Chemistry
7. __________________ – Displays output of the detection

system
SINGLE BEAM SPECTROPHOTOMETER
 Simplest Type; Designed to make one measurement at a
time at one specified wavelength
Westgard Errors on LJ Chart

Table 3. Westgard Control Rules
Random Errors Systematic Errors
_______ – 1 control value _______ – 2 consecutive
exceeds ±2SD; rejection or control values exceed Components of a single-beam spectrophotometer.
warning rule either ±2SD A, Exciter lamp; B, entrance slit; C, monochromator; D,
_______ – 1 control value _______ – 4 consecutive exit slit; E, cuvet; F, photodetector; G, light-emitting
exceeds ±3SD control values exceed ±1SD diode (LED) display
_______ – Range/ _______ – 10 consecutive DOUBLE BEAM SPECTROPHOTOMETER
difference between the control values fall on 1 side  Splits monochromatic light into two components: one
highest and lowest control or the other side of the beam passes through the sample and the other through
result within an analytical mean a reference solution or blank
run is 4SD 1. ___________________________ – 2 photodetectors
AUTOMATION
Automation
 Wavelength – distance between two successive peaks
 400-700 nm – visible spectrum
 <400 nm – ultraviolet region (UV)
 >700 nm – infrared region
 Didymium or holmium oxide filter is used to check
wavelength accuracy
 Neutral density filters and dichromate solution verify _______________________. A, Exciter lamp; B,
absorbance accuracy mirror; C, entrance slits; D, monochromators; E, exit
 Beer-Lambert’s law slits; F, cuvets; G, photodetectors; H, light-emitting
 A = abc = 2 – log%T diode (LED).
o A: molar absorptivity 2. ____________________________ – 1 photodetector
o B: length of light through the solution and 1 chopper or rotating sector mirror
o C: concentration of absorbing molecules
o T: transmittance
 One-point calcuation or calibration
𝐶𝑜𝑛. 𝑜𝑓 𝑆𝑡𝑎𝑛𝑑𝑎𝑟𝑑 (𝐶𝑠) 𝐶𝑜𝑛𝑐. 𝑜𝑓 𝑢𝑛𝑘𝑛𝑜𝑤𝑛 (𝐶𝑢)
=
𝐴𝑏𝑠. 𝑜𝑓 𝑆𝑡𝑎𝑛𝑑𝑎𝑟𝑑 (𝐴𝑠) 𝐴𝑏𝑠. 𝑜𝑓 𝑢𝑛𝑘𝑛𝑜𝑤𝑛 (𝐴𝑢)
SPECTROPHOTOMETRY
 Measurement of light transmitted by a solution to

determine the concentration
PARTS OF A SPECTROPHOTOMETER
1. __________________ – Provide Polychromatic light
___________________________
2. __________________- Minimizes unwanted or stray
light; prevents entrance of scattered light
3. __________________ – Isolates specific or individual
wavelength of light  Excitation of electrons from lower to higher energy state
4. __________________– Controls the width of light beam  Measures light emitted by single atom burned in flame;
(bandpass) measures excited ions (Na+ and K+)
5. __________________ – Holds the solution whose
concentration is to be measured
6. __________________ – Detects and converts transmitted  Element is not excited but merely dissociated from its
light into photoelectric energy chemical bond and placed in an unionized, unexcited
ground state

Clinical Chemistry
 Measures light absorbed by atoms dissociated by heat; GAS CHROMATOGRAPHY

measures unexcited trace metals (Ca2+ and Mg2+)  for naturally volatile compounds or easily converted to
volatile form
 ___________________ – based on fragmentation
and ionization of molecules using a suitable energy
 Unknown sample is made to react with known solution
source
in the presence of an indicator  _______________ – gold standard for drug testing
 Schales and Schales (Chloride)  _______________________ – detects 20 inborn
 EDTA titration (Calcium) errors of metabolism from a single blood spot
LIQUID CHROMATOGRAPHY
TURBIDIMETRY
 based on distribution of solutes between a liquid mobile
 Determines the amount of light blocked by a particulate
phase and a stationary phase
matter in a turbid solution
 High performance liquid chromatography (HPLC) –
 Used in measuring proteins and bacterial suspensions used in rapid HbA1c testing
 Liquid chromatography-Mass Spectroscopy (LC-MS)
NEPHELOMETRY – used in detecting non-volatile substances;
complementary to GC-MS
 Determines amount of scattered light by a particulate
matter in a turbid solution FLUOROMETRY/MOLECULAR LUMINESCENCE
 Used in measuring antigen-antibody complexes SPECTROPHOTOMETRY
 Determines the amount of light emitted by a molecule

after excitation by electromagnetic radiation
 Uses 2 monochromators; measures amount of light
intensity present over a zero background; affected by
quenching
Optical Arrangements of Nephelometry and

Turbidimetry
ELECTROPHORESIS
 Migration of charged particles in an electric field

 Separates proteins on the basis of electrical charge;
Buffer: Veronal/Barbital (pH 8.6) Components of a Fluorometer
DENSITOMETRY
 Measures absorbance of stain  Chemical reaction yields electronically excited

 Scans and quantitates electrophoretic pattern; compound that emits light as it returns to its ground
measures concentration of dye and protein fraction state
 Emission of light is created from a chemical or
ISOELECTRIC FOCUSING electrochemical reaction; usually used in immunoassays
 Migration through a pH gradient
 *pH gradient – created by adding acid to anodic area OSMOMETRY
and base to the cathode area
 Based on measuring changes in colligative properties of
 Ideal for separating proteins of identical sizes but with solutions
different net charges; detects CSF oligoclonal banding
 Freezing-point depression osmometry – most commonly
used method
CHROMATOGRAPHY
 Separation of soluble components based on physical

and chemical characteristics

Clinical Chemistry
ELECTROCHEMISTRY TECHNIQUES CARBOHYDRATES

GLUCOSE
 Glucose is the principal and almost exclusive
 Measurement of electrical potential due to free ion
carbohydrate circulating in the blood
activity
 Glucose is the central, pivotal point of carbohydrate
 Use: _______________________
metabolism
 ____________ is the most important glucose consumer.
 Measurement of electricity (Coulombs) at fixed
CNS consumes about 50% of glucose used by the body
potential
 Glucose can be derived from (1) diet, (2) from body
 Use: _______________________
stores like glycogen, and (3) from endogenous synthesis
from proteins or glycerol of triglycerides.
 Measurement of current flow produced by oxidation
reaction
REGULATION OF BLOOD GLUCOSE
 Use: _______________________ CONCENTRATION
PROCESSES INVOLVED IN CARBOHYDRATE METABOLISM
 Measurement of differences in current at constant 1.
voltage  Metabolism of glucose molecule to pyruvate or
 Use: Specific for pO 2 and glucose lactate to energy
 Decreases blood glucose since glucose is consumed
 Measurement of current after which a potential is to produce lactate/pyruvate
applied to an electromechanical cell 2.
 Lead and iron testing (anodic stripping voltammetry)  Formation of glucose-6-phosphate from non-
carbohydrate sources
THREE BASIC APPROACHES  Increases blood glucose; new glucoses are formed
from other sources
3.
 Samples flow through a common reaction vessel; uses a
 Breakdown of glycogen to glucose for use as energy
system of continuous tubing; ____________________
 Increases glucose due to glycogen degradation
 Mixing of Sample and Reagent: Glass coil inserted into
4.
the flow path  Conversion of glucose to glycogen for storage
 Decreases gluceose since excess glucoses in the
 Uses acceleration and deceleration of rotor to transfer body is stored in the liver and skeletal muscle as
reagents and sample from one chamber to another; glycogen
___________________________ 5.
 Mixing of Sample and Reagent:  Conversion of carbohydrates to fatty acids
____________________________________________  Decreases glucose since carbohydrates are
converted into fatty acids and stored as fats
 Uses syringe pipettes (positive-liquid displacement) to 6.
aspirate and dispense samples; most versatile and most  Breakdown of fats; fats are used as energy
popular; ___________________________________ HORMONES INVOLVED
 Mixing of Sample and Reagent: Magnetic driven Teflon 1. Hyperglycemic hormones
stirring bar, etc  Glucagon, Epinephrine, Cortisol, Growth hormone,
 ____________________________ – all samples are Thyroxine
loaded at the same time and a single test is conducted 2. Hypoglycemic hormone
on each sample
 Insulin
 ____________________________ – more than one test
is analyzed concurrently on a given clinical specimen 3. Regulator hormone
 ____________________________ – any test can be  Somatostatin – inhibits release of growth hormone,
performed on any sample in any sequence insulin, and glucagon
 ____________________________ – multiple tests are
analyzed one after another on a given specimen
 ____________________________ – a system other SPECIMEN FOR GLUCOSE DETERMINATION
than the manufacturer’s reagents can be utilized for  Standard clinical specimen is ______________________
measurement  Fasting Blood Sugar should be obtained after
 ____________________________ – a system where the _____________________________________
operator can only use the manufacturer’s reagent
 Venous blood has lower glucose levels compared to
arterial blood

Clinical Chemistry
 Capillary blood has higher glucose levels compared to Table 4. Comparison Between Type 1 and Type 2
venous blood DM
 Whole blood gives approximately 10 – 15 % LOWER TYPE 1 DM TYPE 2 DM
glucose levels than serum or plasma Pathogenesis
 To convert whole blood glucose into serum or plasma
level, multiply value by 1.15 Incidence rate 5-10% 90-95%
 A serum specimen is appropriate for glucose analysis if Onset Any; most Any; most
serum is separated from the cells within 30 – 60 minutes common to common with
 Glucose is metabolized at room temperature at a rate of childhood/teens advancing age,
____________________ race/ethnicity,
 At 4°C, glucose decreases by approximately hypertension,
____________________ dyslipidemia,
 10% contamination with 5% dextrose will increase polycystic
glucose by 500mg/dL or more ovarian
 CSF glucose concentration is approximately syndrome
______________________ that of plasma concentration Risk Factors Genetic, auto- Genetic, obesity,
 Blood glucose should be obtained 1 – 2 hours before the immune sedentary
spinal tap lifestyle,
 CSF for glucose analysis should be performed polycystic
immediately. If delay in measurement is unavoidable, the ovarian
sample must be centrifuged and stored at 4°C or at -20°C syndrome,
dyslipidemia and
CLINICAL SIGNIFICANCE OF GLUCOSE RESULTS hypertension
HYPOGLYCEMIA C-peptide levels
 Glycemic factors such as glucagon are released when

Pre-diabetes
glucose levels reach ___________________________
 Observable signs and symptoms of hypoglycemia appear
Symptomatology Symptoms Symptoms
when glucose levels reach ________________________
develop abruptly develop
 Critical value for glucose is _________________;
gradually (some
excessively low glucose values can cause severe CNS
patients are
dysfunction especially if blood glucose value drops to 20
asymptomatic)
– 30 mg/dL
Ketosis
 Whipple’s Triad: low blood glucose concentration, typical
symptoms and symptoms alleviated by glucose
Medication Insulin absolute Oral agents
administration
HYPERGLYCEMIA
GESTATIONAL DIABETES MELLITUS
Laboratory Finidngs in Hyperglycemia
 A disorder characterized by impaired ability to
1. Increase glucose in plasma and urine
metabolize carbohydrate usually caused by a deficiency
2. Increase in urine specific gravity
3. Ketones in serum and urine of insulin, metabolic or hormonal changes
4. Decreased blood and urine pH (acidosis)  It occurs during pregnancy and disappears after delivery
5. Electrolyte imbalance (decrease Na+ and HCO3+,  Screening should be performed between 24-28 weeks of
increase K+) gestation
DIABETES MELLITUS (DM)  Screening and diagnostic test: 2-hour OGTT using 75g
glucose load
 Group of metabolic disorders characterized by
hyperglycemia resulting from defects in insulin secretion,  Infants born to diabetic mother are at increased risk for
insulin receptors or both respiratory distress syndrome, hypocalcemia and
hyperbilirubinemia
 Fasting plasma glucose concentration ≥126 mg/dl on
more than one testing is diagnostic of hypoglycemia  After giving birth, women with GDM should be evaluated
6-12 weeks postpartum
 Glucosuria occurs when the plasma glucose levels exceed
________________________ with normal renal function  GDM converts to DM within 10 years in 30-40% of cases
 Severe DM, the ratio of β-hydroxybutyrate to  Diagnostic Criteria for GDM

acetoacetate is 6:1 1. FBS - ≥92 mg/dL
2. 1-hour OGTT = ≥180 mg/dL
3. 2-hour OGTT = ≥153 mg/dL

Clinical Chemistry
GLUCOSE METHODOLOGIES  Guidelines for OGTT

CHEMICAL METHODS o Patient is aked to consume a normal to high
carbohydrate intake (150g carbs per day) for 3
OXIDATION REDUCTION METHOD
days prior to the test
1. _____________________________ – reduction of
cupric to cuprous ions forming cuprous oxide in hot o Patient is asked to fast overnight and to avoid
alkaline solution excessive physical activity. Patient should fast at
 Folin Wu Method least ____________ but not greater than 16 hrs
 Nelson Somogyi Method o OGT testing should be performed on the
 Neocuproine Method morning to prevent hormonal diurnal effect on
 Benedict’s Method (Modified Folin Wu) – glucose.
uses citrate or tartrate as stabilizing agent o Patient should be _______________________.
2. ________________________________ - reduction of Patient should refrain from exercise, eating, or
yellow ferricyanide to colorless ferricyanide (inverse drinking (except water) and smoking
colorimetry) o FBS is measured before giving the glucose load;
CONDENSATION METHOD if the FBS is greater than 140, test should be
 AKA Ortho-toluidine/Dubowski method terminated; if the FBS is <140 mg/dL, glucose
load should be given to the patient.
ENZYMATIC METHODS o Glucose load for an adult is ____ and the patient
should finish drinking it within ______________
o Patient should not vomit, if they vomit,
 measures ___________________________; also
discontinue the test
measures glucose in CSF and urine; presence of bleach
o Blood Glucose is taken every 30 minutes for 2
can cause a false increase in glucose
hrs
 Colorimetric (Saifer Gernstenfield) - enzymes used:
 Intravenous Glucose Tolerance Test (IVGTT)
glucose oxidase, peroxidase
o Used for patients with GI disorders (eg.
 Polarographic - measures the rate of oxygen
Malabsorption)
consumption which is proportional to glucose
o 0.5 g of glucose/kg body weight (given w/in 3
concentration using an oxygen-sensing electrode;
mins) is administered intravenously
enzymes used: glucose oxidase, catalase
o Second blood collection is 5 minutes after
infusion
 Most specific glucose method; reference method; uses
2-HRS POSTPRANDIAL BLOOD SUGAR (2HPPBS)
__________________________________ which is the
 measures how well the used in the diagnosis of GDM
most specific enzyme/reagent in glucose measurement;
 FBS is measured initially, patient is then given glucose
not affected by ascorbic acid or uric acid
load (75g) and plasma glucose is determeined after 2 hrs
 enzymes used: _________________________________
 Normally, blood glucose levels should be back near the
________________
reference limits approximately 2 hrs post load
GLUCOSE DEHYDROGENASE
GLYCOSYLATED HEMOGLOBIN (HbA1c)
 glucose is reduced to a chromophore that is measured
 Reliable method in monitoring long-term glucose control
spectrophotometrically
 Reflects average blood glucose level over the previous 2-
 Mutarotase: shorten time necessary to reach equilibrium
4 months
 enzymes used: glucose dehydrogenase and diaphorase
 Specimen: __________________________
 Methods: Electrophoresis, Immunoassay, HPLC, and
LABORATORY TESTS
Affinity Chromatography
 For every _____________ change in HbA1c, 35 mg/dL is
 Blood Glucose taken any time of the day and without any
added to plasma glucose
fasting
Table 5. Glucose Test Categories in mg/dL
 Requested during insulin shock and hyperglycemic
NORMAL IMPAIRED/ DIAGNOSTIC
ketonic coma
HIGH RISK FOR DM
FBS
 Measure of over-all glucose homeostasis
OGTT
 Requirement: non-per orem (NPO) at least 8 hours
HbA1c (%)
before the test
 Short term glucose control (3 – 6 weeks)

 determines how well the body metabolizes glucose; used
in the diagnosis of GDM  Monitoring DM individuals with Chronic Hemolytic
Anemia and Hb variants

Clinical Chemistry
VALUES TO REMEMBER o Catalyzes the esterification of cholesterol (HDL)

 Fasting Plasma Glucose: resulting in the formation of lysolecithin and
 Glycosylated Hgb: 4-6% (Henry’s); 4.5-8% (Bishop) cholesterol ester
 Conversion Factor:  ____________________________ (30%)
 Critical Values: <40 mg/dL and >500 mg/dL  Found in plasma, serum and rbcs
 Free cholesterol and phospholipids are found on the
LIPIDS surface of lipoproteins
 Are insoluble in blood and water but soluble in organic
solvents (chloroform and ether) TRIGLYCERIDE (TAG)
FORMS OF LIPIDS  Contains 3 molecules of fatty acid and one molecule of
FATTY ACID glycerol by ester bonds; main storage lipid in man
 Linear chains of carbon-hydrogen bonds that terminate  Very hydrophobic and water insoluble – does not contain
with a carboxyl group charged or hydrophilic groups
 Mostly found as constituents of phospholipids or  Evaluates suspected atherosclerosis and measure’s the
triglycerides body’s ability to metabolize fat
 Only a small amount is present in the plasma (free  TAG and cholesterol are the most important lipids in the
unesterified form), most is bound to albumin management of coronary artery disease
 Very important sources of energy  Prior to venipuncture, ideally patients should undergo
 Provide the substance for conversion to glucose fasting for ________________________
(gluconeogenesis)
LIPOPROTEINS
PHOSPHOLIPID  Are large macromolecular complexes of lipids with
 Most abundant lipid derived from phosphatidic acid specialized proteins known as apolipoproteins
 Amphipathic: with polar hydrophilic (water-loving) head  Main purpose: transport TAG and cholesterol to sites of
groups and nonpolar hydrophobic (water-fearing) fatty energy storage and utilization
acid side chains  Apolipoprotein: helps keep the lipids in solution during
 Similar in structure in triglycerides, except they contain circulation through the blood stream; contain a
two fatty acids structural motif called an “amphiphatic helix”
PHOSPHOLIPIDS IN THE BODY MAJOR LIPOPROTEINS
 Phosphatidylcholine (70 – 75%) 1.
 Sphingomyelin (18 – 20%)  Largest and lightest among the lipoproteins;
 Phosphatidylserine, Phosphatidylethanolamine (3 – lipoprotein with LDL
6%)  80-95% TAG by weight
 Transports _________________________
 Lysophosphatidylcholine (4 – 9%)
 Causes non-fasting lipemia
 No charge; remains in the origin during
CHOLESTEROL electrophoresis
 Found on the surface of lipid layers and synthesized in 2.
the liver  Pre-beta Lipoprotein
 Transports __________________________
 Precursor of 5 major classes of steroids: progestins,
3.
glucocorticoids, mineralocortocoids, androgens and
 AKA as the bad cholesterol; beta lipoprotein
estrogens
 Transports cholesterol from
 Evaluates the risk for atherosclerosis, myocardial and __________________________________________
coronary artery occlusions  Transport majority (75%) of the cholesterol
 Direct relationship between elevated serum cholesterol  Directly proportional to the risk of atherosclerosis
and myocardial infarction and coronary heart disease (CHD); higher LDL means
FORMS OF CHOLESTEROL higher risk
 _________________________________ (70%) 4.
 Found in plasma and serum; cholesterol bound in  Smallest and heaviest among the lipoproteins
 Fastest towards the anode; alpha lipoprotein; good
fatty acid
cholesterol
 Neutral lipid: located at center of lipid drops
 Inversely proportional to the risk of atherosclerosis
 Undergoes esterification by lcat
and CHD; lower HDL means higher risk
 Lecithin-Cholesterol Acyl Transferase (LCAT)  Responsible for reverse cholesterol transport;
transports cholesterol ________________________

Clinical Chemistry
MINOR LIPOPROTEINS ENZYMATIC METHODS

1.  Most common method of quantifying the cholesterol
 Product of VLDL catabolism – VLDL remnant; oxidase reaction is to measure the amount of hydrogen
“subclass of LDL” peroxide produced
 Major apolipoprotein: 1. Cholesterol Oxidase Method (Spectrophotometric)
 Density: 1.006-1.019 kg/L = floats on the 1.063  Enzymes used: Cholesterol esterase, Cholesterol
density potassium bromide solution oxidase, Peroxidase
2. TRIGYLCERIDE
 Known as ________________________________
due to electrophoretic mobility same as VLDL but CHEMICAL METHODS
density like LDL 1.
 LDL-like particles that have a molecule of Apo (a)  Uses chromotrophic acid
linked to Apo B-100 by a disulfide bond  End product: Blue colored compound
 Independent risk factor for atherosclerosis  __________________________________________
 Density: 1.045-1.080 kg/L (CDC Reference method)
o alkaline hydrolysis (saponification) using
ABNORMAL LIPOPROTEINS alcoholic KOH, solvent extraction with
1. chloroform and the extract is treated with silicic
 Abnormal lipoprotein found in obstructive jaundice acid (chromatography) to isolate TAG and a
and LCAT deficiency color reaction with chromotropic acid giving rise
 A specific and sensitive indicator of cholestasis to a pink end color
 Lipid content is mostly phospholipid and free 2.
cholesterol (90%); contains ApoC and albumin  Uses diacetyl acetone and ammonia
2.  End product: Diacetyl lutidine compound
 Known as “___________________________ – ENZYMATIC METHODS
density of VLDL during ltracentrifugation but  Specific, Rapid, and easy to use
migrates with LDL in the β-region during  Major Interference: Endogenous Glycerol
electrophoresis
 Glycerol Kinase
 Found in type 3 hyperlipoproteinemia or
 Reaction A - lipase, glycerol kinase, pyruvate kinase
dysbetalipoproteinemia
and LDH
 Also known as VLDL rich in cholesterol due to
 Reaction B - lipase, glycerol kinase, glycerol PO 4
defective catabolism of VLDL
dehydrogenase, diaphorase
METHODOLOGIES LIPOPROTEIN
CHOLESTEROL 1. Ultracentrifugation
CHEMICAL METHODS  Reference method for quantification of lipoproteins
 Based on protein and triglyceride contents of
 PRINCIPLE: Dehydration and oxidation of cholesterol to
lipoproteins
form a colored compound
 Expressed on svedverg units
1.
________________________ – end product:  Reagent: potassium bromide solution (1.063)
Cholestadienyl monosulfonic acid (green end color) 2. Electrophoresis
2. ________________________ - end product:  Electrophoretic pattern (from origin): Chylomicrons,
cholestadienly disulfonic acid (red end color) LDL, VLDL, HDL
 Color Developer Mixture (Liebermann Burchardt  Preferred supporting medium: agarose gel
reagent)  Lipid staining dyes: Oil red O, Fat red 7B, Sudan Black
 Glacial acetic acid B
 Acetic Anhydride 3. Chemical Precipitation
 Concentrated H2SO4 a. HDL
Table 6. General Methods o Uses dextran sulfate (synthetic heparin) with
METHOD STEPS OTHER NAME magnesium (precipitants)
Pearson, Stern o CDC reference method: ultracentrifugation,
One-step Colorimetry
and MacGavack heparin manganese precipitation and Abell-
Two-step Extraction + Colorimetry Bloors Kendall assay
Saponification + o Homogeneous assays are the most popular
Three-step Abell-Kendall method for measuring HDL-C
Extraction + Colorimetry
Saponification + b. LDL
Schoenheimer i. β-quantification
Four-step Extraction + Colorimetry
Sperry o EDTA plasma is the preferred sample
+ Precipitation

Clinical Chemistry
o Plasma is centrifuged for at least 18 IIb (Familial Combined High LDL & VLDL (High
hours; VLDL and CM accumulate as Hyperlipidemia) Cholesterol & TAG)
floating layer, leaving predominantly LDL III (Familial Presence of β-VLDL, High
and HDL in solution Dysbetalipoproteinemia) Cholesterol & TAG)
ii. Homogeneous Direct LDL-C method IV (Familial Hight VLDL (High TAG)
o First reagent: selectively removes non- Hypertriglyceridemia)
LDL lipoproteins High VLDL and Presence of
o Second reagent: releases cholesterol V CM (High Cholesterol and
from LDL so that it can be measured TAG)
enzymatically LIPID STORAGE DISEASES
4. Chromatographic methods
Table 8. Lipid Storage Diseases
 Uses either Gel Chromatography or Affinity
Chromatography Lipid Storage Disease Enzyme Deficient
5. Immunochemical methods Fabry’s disease Alpha galactosidase
 Uses antibodies specific to epitopes on the GM-1 Gangliosidosis Beta galactosidase
apolipoproteins Gaucher Beta glucosidase
6. Immunoassay or Immunonephelometry Krabbe Cerebroside beta galactosidase
 Based on the measurement of the turbidity created Niemann Pick Sphingomyelinase
by apolipoprotein-antibody complexes
Metachromatic Arylsulfatase A
 Lp(a) is measured by immunoturbidimetric assay
leukodystrophy
Sandhoff Total Hexosamindase (A & B)
CLINICAL SIGNIFICANCE
Tay Sach Hexosaminidase
ARTERIOSCLEROSIS VS ATHEROSCLEROSIS
VALUES TO REMEMBER
 Arteriosclerosis – general term for the thickening and ATP III CLASSIFICATION FOR LDL, TOTAL & HDL
hardening of arteries CHOLESTEROL & TAG VALUES
 Atherosclerosis – Type of arteriosclerosis; Hardening of
Table 9. LDL CHOLESTEROL
the arteries caused by plaques (made up ofCholesterol,
< 100 mg/dL Optimal
Fatty substances, Cellular Waste products, Calcium and
100 – 129 mg/dL Near Optimal/Above Optimal
Fibrin) that build up inside the arteries
130 – 159 mg/dL Borderline High
CORONARY HEART DISEASE (CHD)
160 – 189 mg/dL High
 Broad spectrum of Heart disease resulting from Imparied ≥ 190 mg/dL Very High
coronary blood flow
Table 10. HDL CHOLESTEROL
 Clinical (Non-Laboratory) Risk factors for CHD < 40 mg/dL Low
 Cigarette Smoking
≥ 60 mg/dL High
 Hypertension (BP >140/90 mmHg)
Table 11. TOTAL CHOLESTEROL
 Family history of premature CHD
< 200 mg/dL Desirable
 Age (Men > 45 years; Women > 55 years)
200 – 239 mg/dL Borderline High
 Obesity
≥ 240 mg/dL High
 Diabetes Mellitus
Table 12. TRIGLYCERIDE
 Sedentary lifestyle
< 150 mg/dL Normal
ANALPHALIPOPROTEINEMIA
150 – 199 mg/dL Borderline high
 Aka Tangier Disease; HDL deficiency
200 – 499 mg.dL High
ABETALIPOPROTEINEMIA
≥ 500 mg/dL Very High
 Aka Bassen-Kornzweig syndrome; Deficiency of apoB REFERENCE RANGES
(B48 and B100); notable acanthocytes in peripheral
 Total Cholesterol – 140 – 200 mg/dL
blood smear
 HDL Cholesterol – (M) 29 – 60 mg/dL; (F) 38 – 75 mg/dL
FREDRICKSON AND LEVY’S HYPERLIPOPROTEINEMIA
 LDL Cholesterol – 57 – 130 mg/dL
PHENOTYPES
 Triglycerides – 67 – 157 mg/dL
Table 7. Fredrickson Classification of
CONVERSION FACTORS
Hyperlipoproteinemia
TYPE LPP PATTERN  Cholesterol (mg/dL to mmol/L) – _________________
I (Familial LPL deficiency) High CM (High TAG)  Triglyceride (mg/dL to mmol/L) – _________________
IIa (Familial High LDL (High Cholesterol) FORMULA FOR LDL-C

Hypercholesterolemia)  LDL-C = _______________________________________

Clinical Chemistry
 Friedewald Method (Indirect)

 VLDL (mmol/L) = ____________________________  most abundant protein in fetal serum
 VLDL (mg/dL) = ____________________________  detectable in maternal blood up to the 7th or 8th month
 De Long Method (Indirect) of pregnancy
 VLDL (mmol/L) = ____________________________  increased (maternal blood):
 VLDL (mg/dL) = ____________________________ ______________________________________________
 tumor marker for _______________________________
PROTEINS α1 ACID GLYCOPROTEIN/OROSOMUCOID (AAG)
 Are synthesized in the liver and secreted by the  greatest affinity for progesterone
hepatocyte into the circulation except immunoglobulins  useful diagnostic tool in neonates with bacterial
(plasma cells) infections
 Amphoteric: can bear positive and negative charges α1 ANTI – CHYMOTRYPSIN
because of their acid and basic amino acid compositions  binds and inactivates prostate specific antigen (PSA)
PLASMA PROTEINS  major form of PSA found in human sera; associated with
FRACTION SPECIFIC PROTEINS
Alzheimer’s disease
Prealbumin Prealbumin
GROUP-SPECIFIC COMPONENT (Gc) GLOBULIN
Albumin Albumin
 exhibits affinity with vitamin D and actin
α1-Globulin α1antitrypsin, α fetoprotein, α lipoprotein,
 method: radial immunodiffusion
α1 acid glycoprotein, α1 antichymotrypsin,
ALPHA 2 – GLOBULIN
inter α-trypsin inhibitor, Gc globulin
α2-Globulin Ceruloplasmin, Haptoglobin, α2
macroglobulin  largest major non-immunoglobulin in plasma
β-Globulin Transferrin, Hemopexin, β2 microglobulin,  increases 10x in ______________________
Complement, Fibrinogen, LDL, VLDL, CRP  found principally in the intravascular spaces
γ-Globulin Immunoglobulin, CRP (In some books)
PREALBUMIN (TRANSTHYRETIN)  copper-binding glycoprotein; imparts blue color to
protein
 Transport protein for ____________________________
 marker for Wilson’s disease: deposition of copper in skin,
 Used to detect malnutrition
liver, brain and cornea (Kayser Fleisher rings)
 Landmark to confirm that the specimen is really CSF
 decreased: Wilson’s disease, Menkes’ kinky-hair
syndrome
 Protein present in highest concentration in the plasma
 General transport protein; maintains osmotic pressure  binds free hemoglobin by its α chain
 Sensitive and highly prognostic marker in cases of cystic  prevents the loss of hemoglobin and its constituent iron
fibrosis into the urine
 “_____________________________________” BETA - GLOBULIN
 Normal life span in circulation is 15 – 19 days
β2 - MICROGLOBULIN
 High serum albumin levels are more often associated
 light chain component of the major human leukocyte
with dehydration or prolonged tourniquet application or
antigen (HLA)
specimen evaporation
 found on surface of most nucleated cells; needed in the
 Low serum albumin levels can be related to:
production of CD8 cells
 Inflammation, Hepatic disease, Urinary loss,
Gastrointestinal loss, Protein-Calorie malnutrition,
Burn injury, Edema, and Ascites  Major Component of the β2 globulin fraction
 TERMS:  transports iron to its storage sites

 Analbuminemia – absence of albumin  increased: hemochromatosis (bronze-skin) and IDA
 Bisalbuminemia – presence of 2 bands in the COMPLEMENT
albumin region  one of the natural defense mechanisms that protects the
 Hypoalbuminemia – decreased levels of albumin human body from infection
ΑLPHA 1 - GLOBULIN  Complement C3: most abundant form in serum
 binds heme released by degradation of hemoglobin;

 Major inhibitor of protease activity
helps in diagnosis of early hemolysis
 Deficiency: emphysematous pulmonary disease and
juvenile hepatic cirrhosis

Clinical Chemistry
BETWEEN BETA AND GAMMA  Can be used to determine if a certain body fluid is a
transudate or an exudate
FIBRINOGEN
 No to lipemia and hemolysis
 most abundant of all the coagulation factors
METHODOLOGIES
 serve as long term marker for prognosis of cardiovascular
disease
GAMMA GLOBULIN  reference method but not routinely used
 based on the measurement of the nitrogen content of
IMMUNOGLOBULIN
protein (1g of nitrogen=6.54g of protein)
 synthesized in the plasma cells
 uses serum treated with tungstic acid to form a protein
 IgG: most abundant in plasma and lymph
free filtrate
 IgA: main antibody found in secretions
 reagent: H2SO4
 IgM: first antibody that appears in response to antigenic
 end product: ammonia
stimulation
 IgD: present mostly on the surface of B cells
 IgE: antibody associated with allergic and anaphylactic  most widely used method
reactions  requires at least 2 peptide bonds and an alkaline medium
 Reagents:
 general scavenger molecule; binds to the C- ______________________________________________
polysaccharide of the pneumococcus  Principle: Cupric ions complex the group involved in the
 cardiac marker: early warning for persons at risk for peptide bond forming a violet-colored chelate which is
coronary artery disease proportional to the number of peptide bonds present
 inflammatory marker: reflect severity of CHD and reflects the total protein level at 545nm.
FOLIN – CIOCALTEU (LOWRY)
 rapid test for presumptive diagnosis of bacterial versus
viral infection  has the highest analytical sensitivity
MISCELLANEOUS PROTEINS  Principle: Oxidation of phenolic compounds such as
tyrosine, tryptophan, and histidine to give a deep blue
 small heme protein found in skeletal and cardiac muscles color
that transports and stores oxygen from hemoglobin to  Main reagent: Phosphotungstic molybdic acid or phenol
intracellular respiratory enzymes of contractile cells reagent
 higher affinity for oxygen than does hemoglobin  Color enhancer: Biuret reagent
 marker for chest pain (angina) and early detection of ULTRAVIOLET ABSORPTION
acute myorcardial infarction (AMI)  Principle: The absorbance of proteins at 210nm due to
 Rises at 1 – 3 hrs; Peaks at 5 – 12 hrs; Returns to normal the absorbance of peptide bonds at that specific
in 18 – 30 hrs wavelength
TROPONINS  All proteins have absorbance at 210 except tryptophan,
 are regulators of actin and myosin tyrosine and phenylalanine (280nm)
 most important marker for cardiac injury REFRACTOMETRY
 alternative test to chemical analysis of serum proteins

 useful for assessment of early and late AMI SALT FRACTIONATION
 sensitive marker for the diagnosis of unstable angina
 Globulins can be separated from albumin by salting-out
 Rises within 3 – 4 hrs; Peak in 10 – 24 hrs; Remains
procedures using sodium salts
elevated for up to 10 – 14 days
 Reagent: sodium sulfate salts
Table 13. Solubility Property of Proteins
 only found in myocardium, greater cardiac specificity
than TnT; highly specific for AMI PROTEIN SOLUBLE INSOLUBLE
 Rises within 3 – 6 hrs; Peak in 14 – 20 hrs; Returns to Albumin Water Saturated Salt Soln
normal in 5 – 10 days Conc. Salt Soln Highly conc salt soln
Hydrocarbon solvents
Globulin Weak Salt Soln Water
 A cardiac marker; diagnostic for congestive heart failure
Hydrocarbon solvents Saturated Salt soln
SPECIMEN CONSIDERATIONS AND PATIENT PREP Conc salt soln
 Serum is preferred; 24-hr urine and serous fluids can also
be used
 Protein in CSF is ________________________ compared
to Plasma Protein

Clinical Chemistry
SERUM PROTEIN ELECTROPHORESIS  when the substrate concentration reaches a

maximal value, higher concentration of substrate no
 The single most significant clinical application of SPE is
longer result in increased rate of reaction
for the identification of monoclonal spike of
3. Cofactors
immunoglobulins and differentiating them from
 nonprotein entities that bind to enzymes before a
polyclonal hypergammaglobulinemia
reaction occurs
 “blip” in the late α2 or early β zone region: free
 Coenzymes – is an organic compound, which when
hemoglobin
increased in concentration, will result to an increase
 small spikes in the β region: iron deficiency anemia
in velocity of the reaction
(transferrin)
 Activators – organic ions that alter the spatial
 polyclonal gammopathy: rheumatoid arthritis and
configuration of the enzyme for proper substrate
malignancy
binding
ABNORMAL SPE PATTERNS
 Metalloenzymes – an organic ion attached to a
1. Gamma Spike – ___________________________ molecule
2. β-γ bridging – ___________________________
4. Inhibitors
3. α2 globulin spike – ___________________________

4. α1 globulin flat curve – ___________________________
o Physically binds to the active site of an enzyme
5. α1, α2 and β globulin spikes - ______________________
and competes with the substrate for that active
site
o The effect of the inhibitor can be counteracted
by adding excess substrate to bind the enzyme

o does not compete with the substrate but look
for areas other than the active site (allosteric
site)
o increasing substrate concentration does not
Abnormal SPE Patterns
reverse the inhibition
DYES

1. ___________________________ – most sensitive, o binds to the enzyme substrate complex
specific, and precise among the dye-binding assays o increasing substrate concentration results to
2. ___________________________ – most commonly increased inhibition
used, measure absorbance 5. Isoenzymes - have the same catalytic reactions but
3. Tetrabromphenol blue – used in urine reagent strip, slightly different molecular structures
sensitive to albumin 6. Temperature
4. Ninhydrin – for amino acids  enzymes are active at 25°C, 30°C or 37°C
5. Methyl orange
 37°C = optimum temperature
6. Hydroxybenzeneazobenzoic acid (HABA)
 45-50°C = enzyme start to denature
7. Coomassie Brilliant Blue
 60-65°C = inactivation of enzymes
8. Pyrogallol Red – used for analysis of fluids with lower
protein concentrations such as urine and CSF 7. pH - most physiologic reactions occur in the pH range of
7 to 8.
VALUES TO REMEMBER
8. Storage - low temperatures (refrigeration/freezing)
 Reference Range:
render enzymes visibly inactive
 Total Protein: 6.5 – 8.3 g/dL
 2° to 8°C = ideal storage temperature for substrates
 Albumin: 3.5 – 5.5 g/dL
and coenzymes
 Globulin = Total Protein – Albumin
 -20°C = preservation for longer periods of time
 Conversion Factor: g/dL to g/L = 10
 room temperature = ideal storage for LDH (LD4 and
LD5)
ENZYMES 9. Hemolysis - most increases enzyme concentration
FACTORS AFFECTING ENZYMATIC REACTIONS 10. Lactescence or Milky serum - decreases enzyme
1. Enzyme Concentration concentration
 The higher the enzyme concentration, the faster is ENZYME NOMENCLATURE
the reaction  1st digit = classification
2. Substrate concentration  2nd and 3rd digits = subclass
 with the amount of enzyme exceeding the amount  4th and 5th digits = serial number
of substrate, the reaction rate steadily increases as
more substrate is added

Clinical Chemistry
Table 14. Classification of Enzymes

CLASS FUNCTION EXAMPLE ENZYMATIC REACTION
Oxidoreductases Catalyze the LDH, MDH, 1. _____________________________ – reaction rate
(-dehydrogenase) removal or addition ICD, G6PD depends only on enzyme concentration
of electrons 2. _____________________________ – reaction rate is
Transferases Catalyze the CK, AST, ALT, directly proportional to substrate concentration
(-transferase, transfer of chemical OCT GENERAL METHODS TO MEASURE ENZYMATIC
-kinase) group other than REACTIONS
hydrogen from one 1. Fixed time/endpoint – reaction proceeds for a
substrate to another designated time, the reaction is stopped and a
Hydrolases Catalyze hydrolysis Esterases – measurement is made.
or splitting of a bond ACP, ALP, CHS, 2. Continuous Monitoring/ kinetic assay – multiple
LPS measurements are made during the reaction; preferred
by the addition of
Peptidases –
water (hydrolytic than fixed-time
Trypsin,
reactions) pepsin, LAP ALKALINE PHOSPHATASE (ALP)
Glycosidase – CHARACTERISTICS
AMS,  Catalyzes hydrolysis of phosphomonoesters (or organic
galactosidases phosphate esters) into alcohol and phosphate at an
Lyase Catalyze removal of Glulatamate alkaline pH (9.0 – 10.0)
(-decarboxylase) groups from decarboxylase,  Requires activator zinc
substrates without pyruvate ISOENZYMES
hydrolysis decarboxylase,
tryptophan  Normal Isoenzymes: Intestinal, Placental, Bone, and Liver
decarboxylase,  Liver and Bone ALP are the most predominant fractions
aldolase  Can be differentiated using electrophoresis, heat
Isomerase Catalyze the Glucose denaturation, and chemical inhibition
intramolecular phosphate  Carcinoplacental isoenzymes include Regan, Nagao, and
arrangement of the isomerase, Kasahara. They are usually found in patients with
substrate ribose malignancy and their characteristics resemble that of
compound phosphate placental ALP
isomerase ELECTROPHORESIS
Ligase Catalyze the joining Synthase  Origin towards anode:
of two substrate 
molecules, coupled  Liver and bone fractions are difficult to resolve during
with breaking of the electrophoresis
pyrophosphate  To improve separation of bone and liver forms, use
bond in ATP or neuraminidase & wheat germ lectin
similar compounds HEAT DENATURATION
GENERAL PROPERTIES OF ENZYMES
 Most Heat Stable to Most Heat Labile
1. Active site - is a water-free activity, where the substrate 
interacts with particular charged amino acid residues
 Heat stability is determined by heating serum at 56C for
2. Allosteric site - is a cavity other than the active site
10 – 15 minutes
 When bound tightly to the enzyme, the coenzyme is
 ___________________________ – most heat stable of
called a prosthetic groups
all the normal ALP isoenzymes
 Apoenzyme (enzyme portion) and coenzyme forms
 Regan ALP – most heat stable among all the types of ALP
a complete and active system known as holoenzyme
CHEMICAL INHIBITION
(apoenzyme + prosthetic group = holoenzyme)
 Digestive enzyme in its inactive form originally  Placental and intestinal ALPs are inhibited by
secreted from the organ of production is called a phenylalanine reagent and 3M urea inhibits bone ALP
proenzyme or zymogen  Levamisole inhibits both liver and bone ALP
ENZYME THEORY
1. Emil Fisher’s (Lock and Key Theory) – the shape of the

key (substrate) must fit into the lock (enzyme)
2. Kochland’s (Induced Fit Theory) – based on the substrate
binding to the active site of the enzyme

Clinical Chemistry
METHODOLOGIES Table 16. Summary of ACP Methods

METHODS SUBSTRATE END
Table 15. Summary of ALP Methods
PRODUCTS
METHODS SUBSTRATE END
Gutman & Phenyl PO4 Inorganic PO4
PRODUCTS
Gutman
Bodansky
Inorganic Shinowara PNPP p-nitrophenol
Shinowara β-
phosphate + Babson, Read, & α-naphthol α-naphthol
Jones glycerophosphate
glycerol Philips phosphate
Reinhart
Roy & Hillman Thymolphthalein Free
King &
Phenylphosphate Phenol MonoPO4 thymolphthalein
Armstrong
Bessy, Lowry, P-nitrophenol or
& Brock Yellow  Elevated in patients with prostatic CA, however it is not
Bowers & Nitrophenoxide specific since it can also be elevated in prostatic
PNPP
McComb Iron hyperplasia and prostatic surgery
Huggins & Phenolphthalein Phenolphthalein  Prostatic ACP (PAP) is used together with prostate
Talalay disphosphate Red specific antigen (PSA) to monitor recurrence of prostatic
α-naphthol cancer
Moss α-naphthol
phosphate  Useful in forensic clinical chemistry – especially in medico
Buffered legal evaluation of rape
Klein, Babson, Free
phenolphthalein  ACP may also be elevated in bone diseases due to
& Read Phenolphthalein
phosphate osteoclastic activity, as well as in Gaucher disease
CLINICAL SIGNIFICANCE ASPARTATE AMINOTRANSFERASE (AST)
CHARACTERISTICS
 Often used in evaluation of Hepatobilliary (obstructive
conditions) and Bone disorders (Osteoblast involvement)  Aka __________________________________________
 Highest elevation of ALP is seen in: Paget’s disease  Involved in the transfer of an amino group between
(Osteitis deformans) aspartate and α-keto acids with the formation of
 Increased in: Hyperparathyroidism, rickets & oxaloacetate and glutamate.
osteomalacia, fractures, and Malignant tumors  2 isoenzymes: cytoplasmic (predominant form in serum)
 Cirrhosis and Hepatitis produce only slight elevations and mitochondrial
 Biliary obstruction (Cholestasis) – 3 -10 times elevation  Major tissue source: cardiac tissue, liver and skeletal
 Physiologic elevation of ALP can be seen in growing muscle
children due to osteoblastic activity CLINICAL SIGNIFICANCE
ACID PHOSPHATASE (ACP)  used in the evaluation of myocardial infarction,
CHARACTERISTICS hepatocellular disorders and skeletal muscle
 catalyzes the same reaction made by ALP, except that it involvement
is active at pH 5.0  Often used in conjunction with ALT for hepatocellular
 tissue sources: __________________ (major source), disorders
RBCs, platelets, liver and bone SPECIMEN CONSIDERATION
ISOENZYMES  Hemolysis should be avoided because it can dramatically
 Electrophoretic Separation increase serum AST concentration
 ____________________ remains in Origin METHODOLOGIES
 ___________________ migrates with great mobility  _______________ – (pH 7.5) uses malate dehydrogenase
 Chemical Inhibition (MD) and monitors the change in absorbance at 340nm
 ___________________ is inhibited by L-tartrate ALANINE AMINOTRANSFERASE (ALT)
 ___________________ is inhibited by 2% CHARACTERISTIC
formaldehyde and 1mM cupric sulfate solution
 catalyzes the transfer of an amino group from alanine to
METHODOLOGIES
α-ketoglutarate with the formation of glutamate and
 ______________________________ = specific pyruvate
substrate; for endpoint reaction
 Major tissue source: _____________________________
 ______________________________ = preferred for
continuous monitoring CLINICAL SIGNIFICANCE
 If not assayed immediately, serum should be frozen to a
 Highest elevation is found in
pH lower than 6.5. With acidification, ACP will be stable
______________________________________________
for 2 days

Clinical Chemistry
 ALT is slightly increased in obstructive jaundice but METHODOLOGIES

markedly increased in necrotic jaundice
 uses ________________________ as the substrate
METHODOLOGIES
 Triolein (purer form of TAG) is also used as a substrate
 Couple Enzymatic reaction: ALT + Lactate for LPS assay
dehydrogenase; pH 7.5, reading at 340nm Table 19. Methods for Lipase
Table 17. Differentiating AST and ALT Determination
AST ALT Methods Principle
Major Organ Cherry Reference method; substrate – 50% olive oil
Affected Crandal Hydrolysis of olive oil after incubation for 24
Substrate hours at 37°C and titration of fatty acids using
NaOH
End products Tietz &
Fiereck
Color developer 2,4 DNPH 2,4 DNPH Peroxidase Most commonly used; do not use 50% olive
Color intensifier 0.4 N NaOH 0.4 N NaOH Coupling oil
Methods Reitman & Reitman & Table 20. Acute Pancreatitis Markers
Frankel Frankel Onset of Peak Duration of
Marker
AMYLASE (AMS) Elevation Activity Elevation
CHARACTERISTICS Amylase 2 – 12 hrs 24 hrs 3 – 5 days
Lipase 6 hrs 24 hrs 7 days
 catalyzes the breakdown of starch and glycogen
LACTATE DEHYDROGENASE
 Isoenzymes: ___________________________________
CHARACTERISTIC
 Major tissue source: _____________________________
______________________________________________  catalyzes the interconversion of lactic and pyruvic acid
CLINICAL SIGNIFICANCE  enzyme virtually found in all cells of the body
 uses NAD+ (nicotinamide dinucleotide) as coenzyme
 ____________________________________________
 __________________ is the major isoenzyme
 Three-fold amylase increase with normal 24 hours urine
 LD-1 is relatively abundant in cardiac muscle; LD-5 is
amylase – repeat serum AMS after polyethylene glycol
more abundant in skeletal muscles
precipitation
 __________________ - alcohol dehydrogenase enzyme;
METHODOLOGIES
responsible for the metabolic conversion of methanol
 STARCH – Substrate for all methods and ethylene glycol
Table 18. Methods for AMS Determination  Conc in Serum: LD-2 > LD-1 > LD-3 > LD-4 > LD-5
Method Principle Table 21. LDH Tissue Sources
Saccharogenic Classic Reference Method; Measures the Isoenzyme Chain Composition Tissue Sources
amount of reducing sugars produced by the LD1 & LD 2
hydrolysis of starch
Amyloclastic Measures AMS activity following the LD3
decrease in substrate concentration
Chromogenic Measures AMS activity by the increase in LD4 & LD5
color intensity of the soluble dye-substrate
solution CLINICAL SIGNIFICANCE
Couple- Measures AMS activity by continuous
 highest serum levels: pernicious anemia and hemolytic
enzyme monitoring technique; Enzymes used: AMS,
disorders
Glucosidase, Hexokinase, G6PD
 10-fold increase in hepatic carcinoma and toxic hepatitis
LIPASE (LPS)
 LD-1 > LD-2 (“flipped pattern”) = seen in myocardial
CHARACTERISTIC
infarction and hemolytic anemia
 hydrolyzes ester linkages of fats to produce alcohol and METHODOLOGIES
fatty acid
Method Principle
 major tissue source: ____________________________
Most commonly used; pH 8.8;
absorbance measured at 340 nm
 ____________________________________ – secreted
exclusively in the pancreas Preferred method;
 rises more slowly compared to amylase

Clinical Chemistry
Table 22. Methods for LDH Determination  increased: skeletal muscle disease, leukemia, hemolytic
CREATINE KINASE (CK) anemia and hepatic cancer
CHARACTERISTIC  Highest level in progressive muscular dystrophy
 catalyzes the transfer of a phosphate group between OTHER CLINICALLY SIGNIFICANT ENZYMES
creatine phosphate and adenosine diphosphate 5’ NUCLEOTIDASE (5’N)
 found in high concentrations only in muscle and brain  marker for hepatobiliary disease and infiltrative lesions
 major tissue sources: brain, smooth and skeletal muscle in the liver
and cardiac muscle
 ___________ – (BB) most anodal and labile isoenzyme;
 catalyzes the transfer of glutamyl groups between
most dominant isoenzyme found in the brain, intestine
peptides or amino acids through linkage at a gamma
and smooth muscle
carboxyl group
 _________ (20%) – (MB) present only in the myocardium
 useful in differentiating the increase in ALP
 _________ – (MM) least anodal; major isoenzyme (94-
 elevated in _________________________________ –
100%)
______________________________
 sensitive indicator of _________________ – most
 a very sensitive indicator of acute myocardial infarction sensitive marker of ____________________________
(AMI) and Duchenne disorder PSEUDOCHOLINESTERASE (PCHE)
 highest elevation of total CK is seen in Duchenne’s
 marker for insecticide/pesticide poisoning
muscular dystrophy (50x)
(organophosphate poisoning) – low serum PchE
Table 23. Acute Myocardial Infarction
 monitor the effect of muscle relaxants (succinylcholine)
Markers
after surgery
Marker Onset of Peak Duration of
ANGIOTENSIN-CONVERTING ENZYME (ACE)
Elevation activity elevation
Myoglobin 1 – 3 hrs 5 -12 hrs 18 – 30 hrs  also known as peptidyldipeptidase A or kininase II
Troponin T 3 -4 hrs 10 – 24 hrs 10 - 14 days  possible indicator of neuronal dysfunction (Alzheimer’s
Troponin I 3 – 6 hrs 12 – 18 hrs 5 – 10 days disease – CSF)
CK – MB 4 – 6 hrs 12 – 24 hrs 48 – 72 hrs  for the diagnosis and monitoring of sarcoidosis
AST 6 – 8 hrs 24 hrs 5 days CERULOPLASMIN
LDH 12 – 24 hrs 48 – 72 hrs 10 – 14 days  marker for Wilson’s disease (hepatolenticular disease)
METHODOLOGIES ORNITHINE CARBAMOYL TRANSFERASE (OCT)
 Adenylate kinase released after red cell lysis interferes  marker for hepatobiliary disease
with CK assay particularly with hemolysis
 Cleland’s reagent and glutathione – partially restore lost NON-PROTEIN NITROGENS
activity of CK UREA
Table 24. Methods for CK Determination CHARACTERISTICS
Methods Principle
 major end product of protein catabolism
pH 9.0; 340 nm
Enzymes used: CK, pyruvate kinase,  constitutes to 45% of the total NPN
lactate dehydrogenase  synthesized in the liver via the Kreb’s Henseleit cycle
 first metabolite to elevate in kidney disease
 Blood Urea Nitrogen (BUN) – pertains to the nitrogen
Most commonly used; pH 6.8; 340 nm
Enzymes used: CK, hexokinase, G-6-PD content only of urea. This value is often obtained using
indirect methods
 Urea Concentration – concentration of urea as a whole
molecule, not just the nitrogen portion. This value is
ALDOLASE
often obtained using the direct methods.
CHARACTERISTIC
METHODOLOGIES
 splits fructose-1,6-diphosphate aldolase into 2 triose
DIRECT METHOD
phosphate molecules in the metabolism of glucose
1.
 ISOENZYMES
 Aka Fearon reaction
 Aldolase A – Skeletal Muscle
 Inexpensive, Lacks specificity
 Aldolase B – WBC, Liver, Kidney
 End product: Yellow Diazine Derivative
 Aldolase C – Brain tissue
2. O-phthaldehyde
 End product: Colored product

Clinical Chemistry
INDIRECT METHOD  Normal BUN to Creatinine Ration = _________________

 Measures urea by converting it first into ammonium ions METHODOLOGIES
using urease; ammonium ions formed from urea are then
CHEMICAL (DIRECT JAFFE)
measured
1.
 Initial Reaction: Urease (derived from jack bean meal)  Sensitive but non-specific method
converts urea to ammonium ions and carbonate ions 2. Lloyd or Fuller’s Earth Method
 Secondary Reactions: Quantifies ammonium ions that  Adsorbent:
form after the initial reaction o ________________ – sodium aluminum silicate
 o ________________ – aluminum magnesium
o Reagents: KI, HgI silicate
o Products measured: Yellow-orange colloid
(Ammonium dimercuric iodide)  Principle: Serum is mixed with alkaline picrate solution
 and the rate of change in absorbance is measured
o Reagents: NaOCl, phenol, sodium nitroprusside between 2 points
o NaOCl chlorinates ammonia into  Jaffe Reagent: Satd picric acid & 10% NaOH
monochloramine; monochloramine reacts with
 popular, inexpensive, rapid and easy to perform
phenol to form indophenol (blue or green)
ENZYMATIC METHOD
o Reaction is maintained at alkaline pH (>10.0)
1. Creatinine aminohydrolase – CK
with sodium nitroprusside acting as catalyst
 enzymes used: creatinine aminohydrolase,
o Product measured: indophenol at 630 – 660 nm
creatinine kinase, pyruvate kinase and lactate

dehydrogenase
o Ammonia + 2-oxoglutarate + NADH + GLDH ->
2. Creatinase – Hydrogen peroxide method
NAD + Glutamate + Water  potential to replace Jaffe method (specific than Jaffe
o Measurement of decrease in absorbance at 340 method)
nm  enzymes: Creatininase (creatinine aminohydrolase),
 creatinase, sarcosine oxidase, peroxidase
o Conversion of unionized urea into ammonium CLINICAL SIGNIFICANCE
and bicarbonate ion; measure increase in
conductivity rate AZOTEMIA
 1. Pre-Renal
 diminished glomerular filtration with normal renal
o Color change; used in dry reagent strips
function
 cause: dehydration, shock, congestive heart failure
DECREASED BUN LEVELS 2. Renal
 Decreased protein intake  damaged within the kidneys
 Liver disease  acute/chronic renal disease, glomerulonephritis
 Vomiting and Diarrhea 3. Post-Renal
 Pregnancy  usually the result of urinary tract obstruction
UREMIA VS AZOTEMIA VALUES TO REMEMBER
 ___________________________ – refers to increase  Reference Value: M = 0.6 – 1.2 mg/dL; F = 0.5 – 1.1 mg/dL
NPNs, particularly urea in blood  Conversion factor: (mg/dL to mmol/L) =
 ___________________________ – increase in NPNs in URIC ACID
blood; defined as increase in NPNs with symptoms of CHARACTERISTIC
organ involvement such as renal failure
 major end product of purine catabolism
VALUES TO REMEMBER
 final breakdown of nucleic acid catabolism in humans
 Reference Value: 7 – 18 mg/dL  formed from xanthine by the action of xanthine oxidase
 Conversion Factor: mg/dL to mmol/L = ____________ in the liver and intestine
 Urea Concentration: BUN x ______________  filtered by the glomerulus but 40% of uric acid is
CREATININE reabsorbed in the kidneys
CHARACTERISTIC METHODOLOGIES
 end product of muscle catabolism CHEMICAL (CARAWAY/HENRY METHOD)
 not affected by protein diet; not easily removed by  Reagent: Phosphotungstic acid (PTA)
dialysis  Reaction: Uric Acid + PTA -> Tungsten blue + Allantoin
 __________________________________; used to
evaluate fetal lung maturity

Clinical Chemistry
 Sodium Cyanide is used to increase color and inhibit  Right lobe is 6x larger than the left lobe
fading  1.2 – 1.5 kg in weight
 Sodium carbonate may be used instead  Smallest functional unit of the liver is known as the
ENZYMATIC hepatic lobule
 _____________________ is the enzyme used  Blood is supplied from two sources: hepatic artery and
 Principle: ______________________________________ portal vein
______________________________________________ FUNCTIONS OF THE LIVER
 Uric acid has a UV absorbance peak at 293 nm Table 25. Functions of the Liver
CLINICAL SIGNIFICANCE Function
HYPERURICEMIA Synthesis Plasma proteins, carbohydrates, lipid,
 Gout – associated with pain and inflammation of the lipoproteins, clotting factors, ketone
joints; (+) birefringent crystals in the synovial fluid bodies, enzymes
 Increased nuclear metabolism – seen in leukemia, Conjugation Bilirubin
lymphoma, multiple myeloma or polycythemia, Detoxification Toxic substances absorbed in the
hemolytic and megaloblastic anemias and Drug intestine and by-products of metabolism
 Chronic renal disease – due to decreased GFR and metabolism
tubular secretion Excretion and Bile acids or salts, bile pigments,
 ______________________ – deficiency in hypoxanthine Secretion cholesterol
guanine phosphoribosyl transferase (HGPT) Storage Fat and water soluble vitamins, glycogen
HYPOURICEMIA Table 26. Liver Function Tests
 Liver disease, defective reabsorption of uric acid by Function Tests
kidneys (i.e. Fanconi Syndrome) Synthesis
VALUES TO REMEMBER
Conjugation
 Reference Value: M = 3.5 – 7.2 mg/dL; F = 2.6 – 6.0 mg/dL
& Excretion
 Conversion Factor: mg/dL to umol/L = ____________
Detoxification
AMMONIA
CHARACTERISTIC
BILIRUBIN
 arises from the deamination of amino acids CHARACTERISTIC
 preferred specimen: arterial blood
 end product of hemoglobin metabolism and principal
METHODOLOGIES
pigment in bile
 Digestion (Kjeldahl) method – nitrogen ion in a protein-  also formed from destruction of heme-containing
free filtrate of the specimen is converted to ammonia proteins such as myoglobin, catalase and cytochrome
using hot concentrated H2SO4 in the presence of catalyst oxidase
 Measurement of ammonia Table 27. Comparison between Conjugated
 Nesslerization reaction – uses Gum Ghatti reagent and Unconjugated Bilirubin
 Berthelot reaction – uses sodium nitroprusside Bilirubin 1 (B1) Bilirubin 2 (B2)
 Glutamate dehydrogenase
 used in the diagnosis of hepatic failure (hepatic coma)

and Reye’s syndrome
 elevated levels of ammonia are neurotoxic and are often
associated with encephalopathy
VALUES TO REMEMBER
 Reference Range: 19 – 60 ug/dL

 Conversion Factor: ug/dL to umol/L = _________
BILIRUBIN METABOLISM AND EXCRETION
1. When RBCs are destroyed in the spleen, hemoglobin is

LIVER FUNCTION TESTS
released
LIVER
2. Hemoglobin is broken down into heme and globin
ANATOMY OF THE LIVER
3. Globin is recycled while heme is divided into iron and the
 Largest and the most versatile organ in the body protoporphyrin ring
 Has two main lobes, separated from each other by 4. Iron is transported into the bone marrow to be used for
falciform ligament RBC sythesis

Clinical Chemistry
5. Protoporphyrin is broken down into biliverdin, and then  Diazo Reagents

further broken down into bilirubin, specifically the  Diazo A = 0.1% sulfanilic acid + HCl
unconjugated form  Diazo B = 0.5% sodium nitrite
6. Unconjugated bilirubin (UB) is transported by albumin  Diazo blank = 1.5% HCl
towards the site of conjugation, the liver
 Principle: Diazo reaction is diazotization of bilirubin to
7. In the liver cells, UB is converted into Conjugated
produce azobilirubin
bilirubin (CB) through the action of the enzyme uridyl
Table 28. Main Bilirubin Methodologies
diphosphate glucoronyl transferase (UDPGT). The
process occurs in the smooth ER of the liver cells. Evelyn Malloy Jendrassik-Grof
8. Conjugated bilirubin goes out of the liver, down to the Accelerator
biliary tree (bile duct) and emptied into the intestine.
9. In the intestine, conjugated bilirubin is converted into pH
urobilinogen. Urobilinogen is a colorless substance Final
formed in the intestine reaction
10. Urobilinogen is converted into urobilin, an orange brown
Maximum
pigment responsible for the natural brown color of the
absorbance
stool. It is then excreted through the stool out of the
body. CLINICAL SIGNIFICANCE
11. Some of the urobilinogen from the intestine is
reabsorbed back into the blood, only to be excreted by  conjugated bilirubin tightly bound to albumin; has longer
the kidneys into the urine.
half-life than other bilirubin
 formed due to prolonged elevation of conjugated
bilirubin in biliary obstruction
 helps in monitoring the decline of serum bilirubin
following surgical removal of gallstones
JAUNDICE
 also called icterus or hyperbilirubinemia; characterized
by yellow discoloration of the skin, sclera and mucous
membranes
 clinically evident when bilirubin levels exceeds 2mg/dl
Table 29. Classifications of Jaundice
Schematic summary of the pathway of bilirubin Urine Urine
(Bili, in brown circles) transport and metabolism Jaundice B1 B2
urobilinogen bilirubin
METHODOLOGIES Pre-Hepatic
 Hemolysis will cause increase bilirubin while lipemia can (Hemolytic
cause decrease bilirubin Anemia)
 Visible icterisia occurs when bilirubin is ______________ Hepatic
 Bilirubin standard solution is usually made up of (Liver
unconjugated bilirubin diseases)
 Conjugated bilirubin produces color in aqueous Post
solution. Hepatic
 Unconjugated bilirubin produces color only after the (Bile duct
addition of alcohol obstruction)
 Total bilirubin is measured 15 minutes after adding DERANGEMNTS IN BILIRUBIN METABOLISM
methanol or caffeine solution 1. _________________________ – Bilirubin transport
 Caffeine-benzoate is preferred over methanol deficit
because the latter promotes protein precipitation  characterized by impaired cellular uptake in
and increases turbidity bilirubin; elevated B1
 _______________________ – most commonly used 2. _____________________________ – Conjugation deficit
method; more sensitive and not affected by hemoglobin  infants are treated with phototherapy; elevated B1
 _______________________
 Sodium acetate = maintains alkalinity
o deficiency of the enzyme UDPGT resulting to
 Sodium tartrate = provides alkalinity total absence of B2
 Ascorbic acid = terminates initial reaction and o (+) kernicterus
destroys excess diazo reagent  _______________________
o partial deficiency of UDPGT
o small amount of B2 is produced

Clinical Chemistry
3. _________________________________ – Bilirubin  Diarrhea, Excess fluid loss

excretion deficit  SIADH, Excess water intake
 blockade of the excretion of bilirubin into the  Adrenal insufficiency
canaliculi caused by hepatocyte membrane defect  Reset osmostat
 elevated B2 and total bilirubin
 Acute or Chronic renal failure
4. __________________________ – Conjugation inhibition
 Nephrotic syndrome, Hepatic cirrhosis, Congestive heart
 familial form of unconjugated bilirubinemia caused
failure
by circulating inhibitor of bilirubin conjugation
 Pseudohyponatremia (hyperglycemia, hyperlipidemia,
 elevated B1
hyperproteinemia)
VALUES TO REMEMBER
VALUES TO REMEMBER
 Total Bilirubin – Direct Bilirubin = Indirect Bilirubin
 Reference Value: 135 – 145 mmol/L
 Reference Values:
 Threshold Critical Value
 Total Bilirubin: 0.2 – 1.0 mg/dL
 Hypernatremia: _________________
 Direct Bilirubin: 0 – 0.2 mg/dL
 Hyponatremia: ________________
 Indirect Bilirubin: 0.2 – 0.8 mg/dL
 Conversion Factor: mEq/L to mmol/L = __________
 Conversion Factor: mg/dL to umol/L = ____________
POTASSIUM
 Critical Value for Bilirubin: > 18 mg/dL
CHARACTERISTIC
 otherwise known as “kalium”

ELECTROLYTES
SODIUM  major intracellular cation – only 2% of the body’s total
CHARACTERISTIC potassium circulates in plasma
 Functions: heart contraction, neuromuscular excitability,
 also known as “natrium”
ICF volume regulation and hydrogen ion concentration
 major extracellular anion hence the major contributor of
METHODOLOGIES
osmolality
 principal osmotic pressure outside the cell; depends 1. Ion selective electrode (Valinomycin gel)
greatly on the intake of excretion of water 2. Flame emission photometry (Violet)
3. Atomic absorption spectrophotometry
HORMONES AFFECTING SODIUM LEVELS
4. Colorimetry (Lockhead & Purcell)
1.
 major electro-regulating hormone
 promotes sodium retention and potassium CAUSES OF HYPERKALEMIA
excretion 1. Decreased Renal Excretion
2.  Acute or Chronic Renal Failure
 endogenous antihypertensive agent; promotes  Hypoaldosteronism; Addison’s disease
natriuresis  Diuretics
 blocks aldosterone and renin secretion and inhibits 2. Increased Intake
the action of angiotensin II and vasopressin  Oral or IV potassium replacement therapy
METHODOLOGIES 3. Cellular Shift
 Acidosis, Muscle/Cellular injury
1. Ion selective electrode (Glass aluminum silicate) – most
 Chemotherapy
commonly used method
 Leukemia (Increased WBC)
2. Flame emission photometry (Yellow)
 Hemolysis
3. Atomic absorption spectrophotometry
4. Increased Intake
4. Colorimetry (Albanese Lein)
 Sample hemolysis, Thrombocytosis
 Prolonged tourniquet use or excessive fist clenching
CAUSES OF HYPERNATREMIA CAUSES OF HYPOKALEMIA
 Diabetes Insipidus 1. GI Loss
 Osmotic dieresis  Vomiting, Diarrhea, Gastric suction, Intestinal
 Loss of thirst tumor, Malabsorption
 Insensible loss of water  Cancer therapy
 Gastrointestinal loss of hypotonic fluid  Large doses of laxatives
 Excess intake of sodium 2. Decreased Intake
CAUSES OF HYPONATREMIA 3. Renal Loss
 Diuretics, Potassium depletion  Diuretics – thiazides, mineralocorticoids
 Aldosterone deficiency, Ketonuria  Nephritis, Renal tubular acidosis,

Hyperaldosteronism; Cushing’s syndrome
 Salt-losing nephropathy, Vomiting

Clinical Chemistry
 Hypomagnesemia CALCIUM
 Acute leukemia CHARACTERISITC
4. Cellular Shift
 present almost exclusively in the plasma
 Alkalosis, Insulin overdose
 involved in blood coagulation, enzyme activity,
VALUES TO REMEMBER
excitability of skeletal and cardiac muscles and
 Reference Value: 3.5 – 5.2 mmol/L maintenance of blood pressure
 Threshold Critical Value  99% is part of the bones and 1% is in the blood and ECF
 Hyperkalemia: _________________ FORMS OF CALCIUM
 Hypokalemia: ________________  Ionized (active) calcium - 50%
 Conversion Factor: mEq/L to mmol/L = _________  Protein-bound calcium - 40%
EFFECTS OF POTASSIUM TO CARDIAC MUSCLES  Complexed with anions - 10%
 8mmol/L – lack of muscle excitability FACTORS AFFECTING SERUM CALCIUM LEVELS
 6-7mmol/L – may alter ECG 
 10mmol/L – fatal (cardiac arrest)  increases intestinal reabsorption of calcium
 3.0-3.4mmol/L – arrhythmia & paralysis  increases reabsorption in the kidneys
CHLORIDE 
CHARACTERISTIC  activates the process of bone resorption
 major extracellular anion – chief counterion of sodium  stimulates the conversion of inactive vit. D to active
 promotes maintenance of water balance and osmotic vit. D3
pressure in conjunction with sodium 
 only anion to serve as enzyme activator  secreted by the parafollicular/C cells of the thyroid
 Functions: maintains osmolality, blood volume and gland
electric neutrality  hypocalcemic hormone – inhibits PTH and vit. D3
 inhibits bone resorption
 Responsible for Chloride shift – an exchange mechanism
between chloride and bicarbonate across the membrane METHODOLOGIES
of RBCs 1. Precipitation and Redox Titration
METHODOLOGIES 
o end product: oxalic acid (violet color)
 Interferences: bromide, cyanide, and cysteine

 Mercurimetric Titration (Schales & Schales)
o end product: chloranilic acid (violet)
 indicator: Diphenylcarbazone
2. Ortho-Cresolpthalein Complexone dyes
 endproduct: Mercuric chloride (blue violet)
 Dye: Arzeno III
 Spectrophotometric method
3. EDTA titration method (Bachra, Dawer & Sobel)
 Mercuric thiocyanate (Whitehorn titration) 4. Ion selective electrode (Liquid-membrane)
 Ferric perchlorate 5. Flame emission photometry
 Colorimetric amperometric titration – Cotlove CLINICAL SIGNIFICANCE
Chloridometer
HYPERCALCEMIA
 ____________________ – most commonly used method
 Primary Hyperparathyroidism (Most Common PTH –
mediated hypercalcemia)
HYPERCHLOREMIA  Familial hypocalciuric hypercalcemia
 Excess loss of bicarbonate  Ectopic secretion of PTH by neoplasms
 Renal tubular acidosis  Malignancy associated (most common non PTH
 Metabolic acidosis mediated hypercalcemia)
HYPOCHLOREMIA  Vitamin D intoxication, Thyrotoxicosis
 Prolonged vomiting, Diabetic ketoacidosis, Aldosterone  Hypoadrenalism, Immobilization with increased bone
deficiency turnover, Milk-alkali syndrome, Sarcoidosis, Multiple
 Salt-losing renal disease (eg Pyelonephritis) Myeloma
 High serum bicarbonate (eg compensated respiratory HYPOCALCEMIA
acidosis or metabolic acidosis)  Primary hypoparathyroidism
VALUES TO REMEMBER  Severe hypomagnesemia
 Longstanding hypercalcemia
 Reference Values: 98 – 107 mmol/L
 Pseudohypoparathyroidism
 Conversion Factor: mEq/L to mmol/L = _________
 Vitamin D deficiency, Chronic renal failure
 Renal tubulopathies, Fanconi’s syndrome

Clinical Chemistry
 Mutations of Vitamin D receptor MAGNESIUM

 Hypoalbuminemia, Acute pancreatitis CHARACTERISTIC
 Rhabdomyolysis  intracellular cation second abundance to potassium; also
 Tetany an enzyme activator
VALUES TO REMEMBER  majority is stored in bones (53%)
 Reference Values:  life threatening symptoms occur if the serum levels
 Total Calcium: 2.15 – 2.5 mmol/L reaches 5mmol/L
 Ionized Calcium: 1.16 – 1.32 mmol/L FORMS OF MAGNESIUM
 Conversion Factor: mg/dL to mmol/L =  2+
Free Mg /Ionized form – 55%
INORGANIC PHOSPHOROUS  Protein-bound Mg2+ - 30%
CHARACTERISTIC  Complexed with ions – 15%
 ______________________ related to calcium; maximally FACTORS AFFECTING MAGNESIUM LEVELS IN BLOOD
absorbed in the _________________________  __________________________________ - increases

 organic phosphorous exists as: renal and intestinal reabsorption of magnesium
 Organic phosphate – principal anion within cells  __________________________________ - increases
 Inorganic phosphate – part of the blood buffer renal excretion of magnesium
 Only inorganic phosphate is measured in clinical METHODOLOGIES
laboratory  Colorimetric methods
FORMS OF PHOSPHOROUS  Calmagite
1. Free/Unbound form – 55%  Formazen Dye
2. Complexes with ions – 35%  Magnesium thymol blue
3. Protein-bound – 10%  __________________________________ – reference
FACTOS AFFECTING PHOSPHATE CONCENTRATIONS method
1. PTH – decreases phosphate by renal excretion  Dye Lake method – Titan yellow dye (Clayton/Thiazole
2. Calcitonin – inhibits bone resorption
yellow)
3. Growth hormone – increase phosphate renal
reabsorption
METHODOLOGIES HYPERMAGNESEMIA
  Acute or Chronic Renal Failure

 most commonly used method to measure inorganic  Hypothyroidism
phosphate  Hypoaldosteronism
 most common reducing agent: pictol  Hypopituitarism
 other reducing agents: elon, ascorbic acid and  Antacids
senidine  Enemas
 endproduct: ammonium-molybdate complex  Cathartics
CLINICAL SIGNIFICANCE  Dehydration
 Bone carcinoma/Bone metastasis
HYPERPHOSPHATEMIA
HYPOMAGNESEMIA
 Renal Failure
 Poor diet; Prolonged magnesium-deficient IV therapy,
 Increased breakdown of cells (eg Intravascular
Chronic alcoholsim
hemolysis)
 Pancreatitis, Vomiting, Diarrhea
 Neoplastic disorders (eg Lymphoblastic leukemia)
 Laxative abuse, Hyperparathyroidism,
 Intensive exercise, Severe infections
Hyperaldosteronism, Hyperthyroidism
HYPOPHOSPHATEMIA
 Hypercalcemia, Diabetic ketoacidosis
 Infusion of dextrose solution
 Diuretics, Excess lactation
 Use of antacids
 Pregnancy
 Alcohol withdrawal
 Tubular disorders, Glomerulonephritis, Pyelonephritis
 Poor diet
 Tetany
 Vomiting
VALUES TO REMEMBER
 Ketoacidosis
VALUES TO REMEMBER  Reference Value: 0.63 – 1.0 mmol/L
 Conversion factor: mEq/L to mmol/L = ____________
 Reference Values: 0.87 – 1.45 mmol/L
 Conversion Factor: mg/dL to mmol/L = _________

Clinical Chemistry
ANION GAP  the pH decreases by 0.015/each Celsius above 37°C

 is the difference between the unmeasured cations
(sodium and potassium) and unmeasured anions
 Evaluate ventilation by lungs
(chloride and bicarbonate)
 is an index or efficiency of gas exchange and not a
 form of quality control for the analyzer used to measure
measure of CO2 concentration in the blood
these analytes
 Increased: renal failure/uremia, ketoacidosis (starvation
or diabetes), poisoning by methanol, ethanol, ethylene  Evaluate metabolic process
glycol or salicylate, lactic acidosis, hypernatremia and  the kidneys regulate pH by excreting acid and
instrument error reabsorption of HCO 3 from the glomerular filtrate
 Decreased: hypoalbuminemia, hypercalcemia,
hyperlipidemia and elevated myeloma proteins  Evaluate degree of oxygenation
 FORMULA:  reflects the availability of the gas in blood but not its
 AG = __________________________________ content
o RV: 8 – 16 mmol/L
Table 30. Levels of Hypoxemia
 AG = __________________________________
pO2
o RV: 10 – 20 mmol/L
Mild 61 – 80 mmHg
Moderate 41 – 60 mmHg
ACID – BASE BALANCE, & VITAMINS Severe ≤ 40 mmHg
ORGANS ASSOCIATED WITH ACID-BASE
CLINIAL SIGNIFICANCE
BALANCE MAINTENANCE
RESPIRATORY ACIDOSIS
 LUNGS
 Help maintain acid-base balance through gas  pH: Decreased; HCO3: Decreased
exchange or respiration  Organ Defective: Lungs
 Rapid or short term compensation  Primary Cause: Hypoventilation
 Analyte(s) controlled: O 2 & CO2  Organ to Compensate: Kidneys
 KIDNEYS  Primary Compensation: Bicarbonate Reabsorption
 Help maintain acid-base balance through  Example: ______________________________________
reabsorption or excretion of bicarbonate _____________________________________________
 Slow but long term compensation RESPIRATORY ALKALOSIS
 Analye controlled: Bicarbonate (HCO3-)
 pH: Increased; HCO3: Increased
BLOOD BUFFERS
 Organ Defective: Lungs
1. Bicarbonate and carbonic acid – major extracellular
 Primary Cause: Hyperventilation
blood buffer
2. Plasma proteins  Organ to Compensate: Kidneys
3. Hemoglobin  Primary Compensation: Bicarbonate Excretion
4. Inorganic phosphate  Example: ______________________________________
HENDERSON-HASSELBACH EQUATION METABOLIC ACIDOSIS
𝑐𝑜𝑛𝑗𝑢𝑔𝑎𝑡𝑒 𝑏𝑎𝑠𝑒
pH = pKa + log  pH: Decreased; HCO3: Decreased
𝑤𝑒𝑎𝑘 𝑎𝑐𝑖𝑑
 Organ Defective: Kidneys
 Primary Cause: Bicarbonate Excretion
 Where
 pKa = 6.1  Organ to Compensate: Lungs
 conjugate base = bicarbonate (HCO 3-)  Primary Compensation: Hyperventilation

 weak acid = carbonic acid (H2CO3)  Example: ______________________________________
METABOLIC ALKALOSIS
𝐻𝐶𝑂3−
pH = 6.1 + log total CO2 = 𝐻𝐶𝑂3− + 𝐻2 𝐶𝑂3
𝐻2 𝐶𝑂3  pH: Increased; HCO3: Increased
𝐻𝐶𝑂3− = total CO2 - 𝐻2 𝐶𝑂3 𝐻2 𝐶𝑂3 = 0.03 x pCO2
 Organ Defective: Kidneys
PARAMETERS  Primary Cause: Bicarbonate Reabsorption
PH  Organ to Compensate: Lungs
 pH of 7.40 is the optimum level for arterial blood  Primary Compensation: Hypoventilation
 the reference range for arterial blood (7.35-7.45) is only  Example: ______________________________________
0.03 pH unit lower for venous blood owing to the SPECIMEN COLLECTION
buffering effects of hemoglobin known as chloride  specimen: arterial blood
isohydric shift  anticoagulant: 0.05ml heparin/ml of blood

Clinical Chemistry
 syringe and needle for arterial blood must be Table 33. Water Soluble Vitamins
preheparinized Chemical Name Deficiency
 use of butterfly infusion sets is not recommended Vitamin B1 Thiamine Infants: Dyspnea,
 liquid form of heparin is not recommended cyanosis, diarrhea,
vomiting
 Samples should be analyzed immediately (<30mins) Adults: Beri-beri,
SPECIMEN CONSIDERATIONS Wernicke-Korsakoff
 On standing: ___________________________________
syndrome (apathy, ataxia
______________________________________________
and visual problems)
 Exposure to air: _________________________________
Vitamin B2 Riboflavin Angular stomatitis,
______________________________________________
dermatitis, photophobia
 Blood samples should be chilled
Vitamin B3 Panthothenic acid Depressed immune
 Glycolysis results to _____________________________ system, muscle weakness
METHODOLOGIES Vitamin B6 Pyridoxine, Infants: Irritability,
GASOMETER seizures, anemia
Pyridoxal
Adult: facial seborrhea
 Van Slyke
Vitamin B12 Cyanocobalamin Megaloblastic anemia,
 Natelson
Neurologic abnormalities
 mercury – to produce vacuum
Vitamin C Ascorbic Acid Scurvy
 caprylic alcohol – anti-foam reagent
 lactic acid Folic Acid Pterolyglutamic Megaloblastic anemia
 NaOH and NaHSO3 acid
ELECTRODES Niacin Nicotinic Pellagra (dermatitis,

acid/Nicotinamide disorientation, weight
 pH (potentiomentry) loss)
 ________________________________________ –
internal reference electrode
ENDOCRINOLOGY
 ________________________________________ –
 Endocrine System: network of ductless glands that
external reference electrode
secrete hormones directly into the blood; regulatory
 pO2 – ________________ (polarography-amperometry)
system of blood
 pCO2 – __________________________ (potentiometry)
 Hormones: are chemical signals produced by specialized
VALUES TO REMEMBER
cells secreted into the blood stream and carried to target
Table 31. Normal Values
tissue; Major function: to maintain the constancy of
Normal Value
chemical composition of extracellular and intracellular
pH
fluids and control metabolism, growth, fertility and
pO2
responses to stress.
pCO2
FEEDBACK MECHANISMS
HCO3-  Positive feedback mechanism –an increase in the
Total CO2 (arterial) product results to elevation of the activity of the system
Total CO2 (venous) and the production rate.
 Negative feedback mechanism –an increase in the
VITAMINS product results to decreased activity of the system and
Table 32. Fat Soluble Vitamins the production rate.
Chemical Name Deficiency CLASSIFICATION OF HORMONES
Vitamin A Retinol Night blindness, growth Table 34. Classification of Hormones
retardation, abnormal Proteins/ Glycoprotein Steroids Amino acid
taste response, Peptides derivative
dermatitis GHRH, CRH, TSH, LH, FSH, Cortisol, Melatonin,
Vitamin E Tocopherols Mild hemolytic anemia TRH, GnRH, hCG, EPO Aldosterone, Serotonin,
(newborn), RBC fragility, Somatostatin, Estrogen, T3, T4,
ataxia PRF, ADH, Progesterone, Epinephrine,
Vitamin D Calciferol Rickets and Osteomalacia oxytocin, GH, Testosterone, Nor-
Vitamin K Phylloquinones, Bleeding disorder, ACTH, PRL, Vitamin D epinephrine
Menaquinones Hemorrhage Calcitonin,
hPL, PTH,
Insulin,
Glucagon

Clinical Chemistry
HYPOTHALAMUS GONADOTROPINS (FSH – FOLLICLE STIMULATING

 above the pituitary gland and is connected to the HORMONE/LH – LUTEINIZING HORMONE)
posterior pituitary by the infundibulum (pituitary stalk)  important markers in diagnosing fertility and menstrual
 supraoptic and paraventricular nuclei produce cycle disorders
vasopressin and oxytocin  ___________________ aids in spermatogenesis in males
HYPOTHALAMIC HORMONES  ____________________________ helps Leydig cells to
1. Thyrotropin-releasing hormone (TRH) produce testosterone (male) and for female, it is
2. Gonadotropin-releasing hormone (GnRH) necessary for ovulation and the final follicular growth
3. Somatostatin/Growth hormone-inhibiting hormone (GH-  _________________________ also acts on thecal cells to
IH) cause synthesis of androgens
4. Growth hormone-releasing hormone (GH-RH) THYROID STIMULATING HORMONE (TSH)
5. Prolactin-inhibiting factor (PIF)  also known as thyrotropin
6. Vasopressin (supraoptic)
 main stimulus for the uptake of iodide
7. Oxytocin (paraventricular nuclei)
 stimulates thyroid hormone synthesis
PINEAL GLAND
ADRENOCORTICOTROPIC HORMONE (ACTH)
 secretes ____________________________________
that decreases pigmentation of the skin  produced in response to low serum cortisol
PITUITARY GLAND  highest levels is between 6:00am to 8:00am; lowest level
 also known as the “master gland” is between 6:00pm to 11:00pm
 located in a small cavity called the sella turcica or Turkish  increased: Addison’s disease, ectopic tumors, after
saddle protein-rich meals
 All pituitary hormones have circadian rhythms  best time for collecting specimen: 8:00am to 10:00am
ADENOHYPOPHYSEAL CELLS PROLACTIN (PRL)
 functions in the initiation and maintenance of lactation
1. ________________________ – secrete growth hormone
2. ______________________________ – secrete prolactin  major inhibitor: _______________________________
3. __________________________________ – secretes TSH  consequence of prolactin excess: hypogonadism
4. ____________________________ – secretes LH and FSH  specimen requirement: blood should be collected 3-4
5. _________________ – secretes proopiomelanocortin hours after the individual has awakened; fasting sample
(cleaved to produce ACTH, β-endorphin and β-lipotropin) POSTERIOR PITUITARY GLAND
ANTERIOR PITUITARY GLAND (ADENOHYPOPHYSIS)
 capable of releasing hormones’ oxytocin and vasopressin
GROWTH HORMONE (GH/SOMATOTROPIN) but not capable of producing it
 most abundant of all pituitary hormones OXYTOCIN
 controlled by GH-RH (the amount released) and  stimulates contraction of the gravid uterus at term –
somatostatin (governs the frequency and duration of “Fergusson reflex”
secretory pulse) ANTI-DIURETIC HORMONE (ADH)/ARGININE VASOPRESSIN
 major stimulus: ________________________________ (AVP)
 Disorders:  acts on the distal convoluted tubule and collecting duct
 Idiopathic GH deficiency – most common cause of  major function: maintain osmotic homeostasis by
pituitary dwarfism in children regulating water balance
 Pituitary adenoma – most common etiology in  promotes reabsorption of water by the renal tubules –
adult-onset GH deficiency maintains water homeostasis
 Acromegaly – due to overproduction of GH  DIABETES INSIPIDUS (DI) is deficiency of ADH; results in
Table 35. Diagnostic Test for Growth severe polyuria
Hormone  Clinical Picture for Diabetes Insipidus
Screening Tests Confirmatory Tests o Normoglycemia, Polyuria with low SG,
GH deficiency Physical activity Insulin tolerance Polydipsia, Polyphagia
(Exercise) test test (gold standard)
 __________________________________________
Arginine stimulation
__________________________________________
test (2nd
o deficiency of ADH with normal ADH receptor
confirmatory test)
 __________________________________________
Acromegaly Somatomedin-C or Glucose suppression
o normal ADH but abnormal ADH receptor – renal
Insulin-like growth test – OGTT (75g
resistance to ADH action
factor glucose)
o failure of the kidneys to respond to normal or
elevated ADH levels

Clinical Chemistry
 Diagnostic test for DI – REIDEL’S THYROIDITIS

__________________________________________  thyroid turns into a woody or stony-hard mass
THYROID GLAND SUBCLINICAL HYPERTHYROIDISM
 also known as the “butterfly-shaped’ gland  shows no clinical symptom but TSH is low and FT 3 and FT4
 consist of two lobes connected by the isthmus located at are normal
the lower part of the neck
 follicle is the fundamental unit of the thyroid gland
 2 types of cells: follicular cells (T3 and T4) and  associated with neck pain, low grade fever and swings in
parafollicular cells/C cells (Calcitonin) thyroid function test
 ________________________________ is a glycoprotein HYPOTHYROIDISM
stored in the follicular colloid of the thyroid gland
 Signs and symptoms: bradycardia, weight gain,
 ____________________________ is the most important
coarsened skin, cold intolerance and mental dullness
element in the biosynthesis of thyroid hormones
PRIMARY HYPOTHYROIDISM
MAJOR THYROID HORMONES
 primarily due to deficiency in elemental iodine
TRIIODOTHYRONINE (T3)  also caused by destruction or ablation of the thyroid
 ______________________________________________ gland
 principal application of this hormone is in the diagnosis 
of T3 thyrotoxicosis  most common cause of primary hypothyroidism
 ______________________________________________  characterized by thyroid replaced by a nest of
 an increase in the plasma level of T 3 is the first lymphoid tissue
abnormality seen in cases of hyperthyroidism  lab result: high TSH and positive TPO antibody
TETROIODOTHYRONINE (T4)  Myxedema
 ______________________________________________  myxedema coma is the severe form of primary
 major fraction of organic iodine hypothyroidism
 the amount of serum T 4 is a good indicator of thyroid  peculiar non-pitting swelling of the skin
secretory rate  clinical features: “puffy” face, weight gain, slow
THYROID-HORMONE BINDING PROTEINS speech, eyebrow thinned, dry and yellow skin and
1. anemia
 transports majority of T3 (affinity for T3 is lower than Table 36. Autoimmune Disorders of the
T4) Thyroid Hormone
 transport 70-75% of total T4 Hashimoto’s Thyroiditis Grave’s Disease
2. Thyroxine-binding prealbumin (transthyretin)
 transports 15-20% of total T4
 T3 has no affinity for prealbumin
3. Thyroxine-binding albumin
 transports T3 and 10% of T4
HYPERTHYROIDISM
 Signs and symptoms: tachycardia, tremors, weight loss, SECONDARY HYPOTHYROIDISM

heat intolerance, emotional lability and menstrual  due to pituitary destruction or pituitary adenoma
changes TERTIARY HYPOTHYROIDISM
THYROTOXICOSIS
 due to hypothalamic disease
 high levels of free thyroid hormones in the circulation
CONGENITAL HYPOTHYROIDISM
 T3 thyrotoxicosis/Plummer’s disease: FT3 increased but
 defect in the development or function of the gland
FT4 normal with low TSH
 symptoms: retarded physical and mental development
 T4 thyrotoxicosis: T3 normal or low but T4 is increased
of the child
with low TSH
Table 37. Clinical Presentation of Patients
with Hypothyroidism & Hyperthyroidism
 most common cause of thyrotoxicosis
Hypothyroidism Hyperthyroidism
 antibodies are produced which are active against the TSH
Basal metabolic
receptor
rate
 features: exophthalmos (bulging eyes) and pretibial
Sympathetic
myxedema
response
 diagnostic test: TSH receptor antibody test
Weight

Clinical Chemistry
Temperature  inversely related to TBG – decreased T3 uptake results to

tolerance elevated TBG result
THYROXINE-BINDING GLOBULIN (TBG)
GI function  used to confirm results of FT 3 and FT4 or abnormalities in
the relationship of the total thyroxine (TT4) and THBR test
 useful in distinguishing between hyperthyroidism
Cardiovascular (elevated T4 + N TBG) and euthyroidism (elevated T 4 +
function elevated TBG)
Table 38. Thyroid Gland Disturbances
T3 & T4 TSH
Respiratory Primary Hypothyroidism
function Secondary Hypothyroidism
General Primary Hyperthyroidism
appearance Secondary Hyperthyroidism
Table 39. Secondary vs Tertiary
Hypothyroidism
General behavior Mental Hypothyroidism Secondary Tertiary
Restlessness
retardation; T3 & T4
Irritability
Mental & TSH
Anxiety
Physical
Hyperkinesis TRH
sluggishness
Wakefulness TRH Stimulation/ TSH before TSH before
Somnolence
Administration administration: administration:
Others Increased Test
Cholesterol & Increased ALP
TSH after TSH after
TAG administration: administration:
THYROID FUNCTION TESTS
TRH STIMULATION TEST PARATHYROID GLAND

 used to differentiate euthyroid and hyperthyroid  located on or near the thyroid capsule; most people have
patients who both had undetectable TSH levels 4 parathyroid hormones
 used to confirm borderline cases and euthyroid Grave’s  smallest endocrine gland; secretes parathyroid hormone
disease – hypercalcemic hormone
PARATHYROID HORMONE
 most important thyroid function test – best method for  to prevent hypocalcemia (regulates blood calcium)
detecting clinically significant thyroid dysfunction  preserve calcium and phosphate within normal range
 most clinically sensitive assay in the detection of primary  promotes bone resorption – release calcium in the
bloodstream
thyroid disorders
 increases renal calcium reabsorption
REVERSE T3 (rT3)  stimulates conversion of inactive vitamin D to activated
 formed from the removal of one iodine from the inner vitamin D3
ring of T4 HYPERPARATHYROIDISM
 rd
3 major circulating thyroid hormone
 Primary Hyperparathyroidism
FREE THYROXINE INDEX (FTI OR T 7)
 physiologic defect lies within the parathyroid gland
 indirectly assesses the level of T 4 in the blood  most common cause of hypercalcemia
 FTI = TT4 x THBR  due to the presence of a functioning parathyroid
𝑇3 𝑈 (%)
 FTI = TT4 x 100 adenoma
TOTAL T3 (TT3), FREE T3 (FT3) AND FREE T4 (FT4)  Secondary hyperparathyroidism
 used to differentiate drug-induced TSH elevation and  develops in response to decreased calcium
hypothyroidism  diffuse hyperplasia of all 4 glands
 the value of TT3 or FT3 is in confirming hyperthyroidism  Tertiary hyperparathyroidism
T3 UPTAKE  phosphate levels are normal to high; calcium
 measures the number of available binding sites of the phosphates precipitate in soft tissues
thyroxine-binding proteins (TBG)
 does not measure the level of T 3 in serum but it reflects
the serum level of TBG

Clinical Chemistry
Table 40. Hyperparathyroidism Levels  signs and symptoms: obesity but with thin extremities
Primary Secondary (“buffalo hump”), hirsutism, hyperglycemia, thinning of
Serum Calcium the skin, poor wound healing, hypertension,
Serum hypercholesterolemia, low WBC count (lymphocytes)
Phosphate  ________________________________ – current
PTH reference method for measuring urinary free cortisol
Urine Calcium  ________________________________ – most sensitive
Urine Phosphate and screening test
VALUES TO REMEMBER Table 43. Diagnostic Tests for Cushing’s
Syndrome
 Calcium level ________________________ = laryngeal
Screening Tests Confirmatory Tests
stridor and tonic clonic seizures
 Calcium level ________________________ = tetany and 24-hour urinary free Low dose dexamethasone
cortisol test suppression test
altered neuromuscular activity
Overnight dexamethasone Midnight plasma cortisol
ADRENAL GLAND suppression test Corticotropin-releasing
 has pyramid-like shape located above the kidneys Midnight salivary cortisol hormone (CRH) stimulation
 composed of distinct but conjoined glands, the outer test test
adrenal cortex (yellow) and inner adrenal medulla (dark
HYPOCORTISOLISM
mahogany)
 screening test: _________________________________
ADRENAL CORTEX
_____________________________________________
 major site of steroid hormone production  confirmatory test: Insulin tolerance test
Table 41. Adrenal Cortex Layers  _______________________________ – gold standard
Layer Function for secondary and tertiary hypocortisolism
Zona glomerulosa (Outer  Overnight metyrapone test – alternative diagnostic or
10%) confirmatory test for seconda
Zona fasciculata (Middle  ry or tertiary adrenal insufficiency
75%)  Primary hypocorticolism (primary adrenal insufficiency)
Zona reticularis (Inner 10%)  due to decreased cortisol production – 90%
destruction of adrenal cortex
 disorder: Addison’s disease – hypotension,
CORTISOL hyponatremia, hyperkalemia, weight loss,
hyperpigmentation
 is the principal glucocorticoid
 Secondary hypocorticolism (secondary adrenal
 mostly bound to glycoprotein – transcortin
insufficiency)
 stimulates gluconeogenesis in the liver resulting in
 due to hypothalamic-pituitary insufficiency with loss
hyperglycemia
of ACTH
 is the only adrenal hormone that inhibits the secretion of
 absence of hyperpigmentation
ACTH
 highest level ___________________________ and
 results in the deficiency of enzymes necessary for the
lowest at night ____________________________
synthesis of cholesterol which will result to decreased
 specimen: serum (red top), urine; blood sample should
plasma cortisol and increased ACTH and androgen levels
be drawn at 8:00 AM
 definitive tests:
 urine free cortisol levels are sensitive indicators of
 17-OHP measurement in amniotic fluid
adrenal hyperfunction (endogenous hypecorticolism) –
 Genotyping cells from chorionic villous sampling –
24 hour urine collection
preferred
Table 42. Cortisol Urinary Metabolites
ALDOSTERONE
Method Reagent
17- Phenylhydrazine  most potent mineralocorticoid (electro-regulating
hydroxycorticosteroid in H2SO4 + hormone)
alcohol  helps regulate water and electrolytes (sodium, chloride
17-ketogenic steroids Meta- and potassium) and blood pressure
dinitrobenzene  acts on renal tubular epithelium to increase retention of
Na+ and Cl- and excretion of K+ and H+
Hypercortisolism (Cushing’s syndrome)  method: RIA and chromatography
 caused primarily by excessive production of cortisol and

ACTH but decreased aldosterone and renin

Clinical Chemistry
PRIMARY HYPERALDOSTERONISM (CONN’S DISEASE) EPINEPHRINE (ADRENALINE/SECONDARY AMINE)

 caused by aldosterone-secreting adrenal adenoma  most abundant medullary hormone
 associated with elevated plasma aldosterone and low  produced from norepinephrine and only from the
plasma renin adrenal gland
Table 44. Tests for Conn’s Disease
 called the “flight or fight” hormone: release in response
Screening Test Confirmatory Test to physiologic (injuries) or psychologic (stress, anxiety)
Plasma aldosterone Saline suppression test threats
concentration/plasma Oral sodium loading test
Fludrocortisone  major metabolite: __________________________:
renin activity ratio major (60%) catecholamine metabolite in the urine
suppression test
 >30 ratio: __________ Captopril challenge test  minor metabolites: metanephrine, normetanephrine
___________________
and homovanillic acid
___________________
DOPAMINE (PRIMARY AMINE)
 >60 ratio: __________
___________________  catecholamine produced via the decarboxylation of 3,4-
___________________ dihydroxyphenylalanine
SECONDARY HYPERALDOSTERONISM  major intact catecholamine in the urine
 occurs as a result of excessive renin production  major metabolite: ___________________________
 accompanied by elevated plasma levels of aldosterone PHEOCHROMOCYTOMA

and renin  tumor of the adrenal medulla or sympathetic ganglia
 associated disorders: Liddle’s syndrome  due to the overproduction of catecholamine
(pseudohypoaldosteronism), Barterr’s syndrome  screening test: plasma metanephrines and
(bumetanide-sensitive chloride channel mutation), normetanephrines by HPLC (four-fold increase)
Gitelman’s syndrome (thiazide-sensitive transporter  diagnostic test: 24-hour urinary excretion of
mutation) metanephrines and normetanephrine
HYPOALDOSTERONISM  pharmacologic test: clonidine test and glucagon
 due to the destruction of the adrenal glands and stimulation test
deficiency of glucocorticoid Patient preparation in the diagnosis of Pheochromocytoma
 also associated with 21-hydroxylase deficiency  Avoid caffeine, nicotine, alcohol and acetaminophen,
 Tests for hypoaldosteronism monoamine oxidase inhibitors and tricyclic
 Furosemide stimulation test or upright posture antidepressants for at least 5 days before testing
 Saline suppression test
NEUROBLASTOMA
WEAK ANDROGENS
 fatal malignant condition in children resulting to
 serve as precursors for the production of more potent excessive production of norepinephrine
androgen and estrogen in tissues
 (+) high urinary excretion of HVA or VMA or both and
 example: dehydroepiandrosterone (DHEA) and dopamine
androstenedione
METHODOLOGIES
 ______________________________: principal adrenal
androgen; converted into estrone  Specimen: _____________________________________
 excessive production of androgens results in virilization  Patient preparation:

(pseudohermaphroditism)  Patient should undergo overnight fasting
 Avoid smoking or drinking coffee at least 4 hours
ADRENAL MEDULLA
prior to blood collection
 composed primarily of chromaffin cells that secrete  Blood is collected on pre-chilled EDTA tubes
catecholamines  Specimen considerations:
 L-tyrosine is the precursor of catecholamines  Catheterization is the preferred method of blood
 ratio of norepinephrine to epinephrine is 9:1 collection
NOREPINEPHRINE (PRIMARY AMINE)  Urine is preserved with 10ml of 6N HCl
 produced by the sympathetic ganglia 1. Chromatography – HPLC or GC-MS (VMA and
 highest concentration is in the brain (CNS) metanephrines)
 major metabolite in CNS is MHPG 2. Radioimmunoassay – sensitive screening test for total
MAJOR METABOLITES OF NOREPINEPHRINE plasma catecholamines
 >2000pg/ml of plasma catecholamines: diagnostic
 3-methoxy-4-hydroxyphenylglycol (MHPG) – CSF and
for pheochromocytoma
urine
 vanillylmandelic acid

Clinical Chemistry
REPRODUCTIVE HORMONES  prime secretory product of the ovary

 testes and ovaries produce sex steroids such as  single best hormone to determine whether ovulation
androgens and estrogens from cholesterol has occurred
 major transport proteins: sex-hormone binding globulin  deficiency: failure of implantation of embryo
 metabolites: pregnanediols (most easily measured
(SHBG), corticosteroid-binding globulin (CBG) and
metabolite), pregnanediones, pregnanalones
albumin
HUMAN CHORIONIC GONADOTROPIN (HCG)
TESTOSTERONE
 produced by the trophoblast cells of the placenta during
 principal androgen hormone in the blood – most potent
pregnancy
male androgen
 same α-subunit as LH, FSH and TSH; similar β-subunit to
 synthesized by the Leydig cells of the testis of the male;
LH – “LH-like” hormone
controlled primarily by the FSH and LH
 intact HCG (α and β) is the predominant form throughout
 function: growth and development of the reproductive
pregnancy
system, prostate and external ganglia
 method: immunometric (sandwich) method – serum and
 tests for male fertility: semen analysis, testosterone, FSH
urine samples
and LH
INHIBIN A
 transport proteins: SHBG (60%) and albumin (40%)
Table 45. Types of Testicular Infertility  Reproductive hormone which inhibits FSH activity
(Hypogonadism) METHODOLOGIES
Testos- FSH & Description/ other
 Porter-Silber – for 17-OHCS
terone LH information
 Zimmerman – steroids with 17-keto structure
Pre-
Due to hypothalamic  Pisano – quantitating metanephrines and
testicular
or pituitary lesions normetanephrines
(Secondary)
 Kober – estrogen
May be congenital
TESTS FOR MENSTRUAL CYCLE DYSFUNCTION
(e.g. Klinefelter’s
Testicular 
syndrome) or
(Primary) TESTS FOR FEMALE INFERTILITY
acquired (e.g.

varicocele)
MARKERS FOR DOWN SYNDROME
Disorders of sperm
Post-  Increased: _____________________________________
transport and
testicular  Decreased: ____________________________________
function
DEHYDROEPIANDROSTERONE (DHEA) GASTRIN
 peptide secreted by the G cells of the antrum of the
 principal androgen formed by the adrenal cortex; weak stomach
androgen  causes secretion of the HCl by parietal cells in the
ESTROGEN stomach
 functions: promotion of breast development,  diagnostic marker for Zollinger-Ellison syndrome (islet-
maturation of the external genitalia, deposition of body cell tumor)
fat and termination of linear growth (secondary sexual SEROTONIN (5-HYDROXYTRYPTAMINE)
characteristics in the female)  derived from the hydroxylation and decarboxylation of
 precursor: acetate, cholesterol, progesterone and tryptophan
testosterone  synthesize by argentaffin cells, primarily in the GI tract
FORMS OF ESTROGEN  diagnostic marker for carcinoid tumor
1. _________________ – most abundant estrogen in post-  tests: Ehrlich’s aldehyde test – (+) purple color
menopausal women
2. _________________ – most potent estrogen; major THERAPEUTIC DRUG MONITORING
estrogen  a quantitative procedure performed for drugs with
 most abundant estrogen in pre-menopausal women narrow therapeutic index
 transport proteins: albumin (60%), SHBG (38%)  the half-life of the drug determines the time to reach the
3. _________________ – estrogen found in maternal urine steady-state or average concentration
 major estrogen secreted by the placenta during  Mixed Function Oxidase (MFO) system: biochemical
pregnancy pathway responsible for the greatest portion of drug
PROGESTERONE metabolism
 produced mainly by the granulose (lutein) cells of the

corpus luteum in the female

Clinical Chemistry
PHARMACOLOGICAL PARAMETERS THAT ANTIBIOTICS

DETERMINE DRUG CONCENTRATION
1. Liberation – release of the drug
2. Absorption – transport of the drug from the site of  Gentamicin, tobramycin, amikacin, kanamycin,
neomycin, streptomycin
administration to the blood
3. Distribution – delivery of the drug to the tissues  Used for treatment of Gram-negative bacterial infection
4. Metabolism – process of chemical modification of the  Toxic effects: nephrotoxocity and ototoxicity
drug by cells
5. Excretion – process by which the drug and its  Used against Gram-positive bacilli and cocci
metabolites are excreted from the body  Toxic effects: “______________________________”,
TERMINOLOGIES nephrotoxicity, ototoxicity
1. Bioavailable fraction (f) – fraction of the dose that CHLORAMPHENICOL
reaches the blood  Toxic effects: blood dyscrasia, cytoplasmic vacuolation
2. First pass hepatic metabolism – drugs that are (erythroid and myeloid cells)
transported to the liver lost a fraction of its bioavailability ANTI-EPILEPTIC DRUGS
before the drug reaches the general circulation
3. Half-life (t1/2) – time required to reduce a drug level to
 Long acting barbiturate that controls grand-mal tonic
half its original value
clonic seizures and focal epilepsies
4. _____________________________ – highest
concentration of a drug obtained in the dosing interval
5. _____________________________ – lowest  Controls tonic-clonic, simple partial seizures; a short
concentration of a drug obtained in the dosing interval term prophylactic agent in brain injury
6. _____________________________ – relationship
between the drug concentration at the target site and  Treatment of petit mal (absence seizures), atomic seizure
response of the tissues and grand mal
7. ______________________ – mathematical expression of
the relationship between drug dose and drug blood level  Effective for grand mal seizures and treating seizures
8. Pharmacogenomics – study of genes that affect the accompanied by pain
performance of a drug in an individual ETHOSUXIMIDE
THERAPEUTIC DRUGS  Drug of choice in controlling petit mal (absence) seizure
CARDIOACTIVE DRUGS
 Class I – rapid Na+ blockers  Chemically similar to neurotransmitter gamma

 Class II – beta receptor blockers aminobutyric acid
 Class III – K+ channel blockers  Adverse effects: dizziness, ataxia, fatigue and nystagmus
 Class IV – Ca2+ channel blockers PSYCHOACTIVE DRUGS
DIGOXIN LITHIUM
 Treatment of atrial arrhythmia and congestive heart  Used for treatment of manic-depressive bipolar
failure (CHF) disorders
LIDOCAINE (XYLOCAINE) TRICYCLIC ANTI-DEPRESSANTS
 Correct ventricular arrhythmia and treatment of MI  Used for treatment of depression, insomnia and extreme
QUINIDINE apathy
 Treatment of arrhythmia  Major metabolite: desipramine
FLUOXETINE (PROZAC)
 Treatment of ventricular arrhythmia  Treatment of obsessive-compulsive disorders
 Toxic effects: reversible lupus-like- syndrome BRONCHODILATOR
DISOPYRAMIDE
 Has anti-cholinergic effects – dry mouth and constipation
 Treatment of asthma and chronic obstructive pulmonary
PROPANOLOL
disease
 Used in treatment of thyrotoxicosis
IMMUNOSUPPRESIVE DRUGS
AMIODARONE
 Iodine-containing drug which can cause hypo or CYCLOSPORINE
hyperthyroidism  For suppression of acute graft-versus-host disease
VERAPAMIL (GVHD)
 Treatment of angina, hypertension and supraventricular  Specimen of choice: whole blood
arrhythmia

Clinical Chemistry
TACROLIMUS  Peak concentrations are drawn one hour after an orally

 100x more powerful than cyclosporine administered dose and immediately after infusion for an
 Toxic effects: thrombus formation, nephrotoxicity, IV administered drug; best specimen
neurotoxicity COLORIMETRY
RAPAMYCIN  Acetaminophen in urine is detected by boiling to form p-
 Similar to tacrolimus with major side effects of amphenol which then reacts with o-cresol to form
thrombocytopenia and lipid abnormalities indophenol blue
MYCOPHENOLATE MOFETIL  Trinder assay for salicylate using ferric nitrate forming a
 Decreases renal allograft rejection colored complex
LEFLUNOMIDE (LFM) IMMUNOASSAY METHODS
 Inhibits lymphocyte proliferation; treatment of RA
 can detect drug levels in nanomolar range; sensitive and
ANTINEOPLASTIC DRUGS
specific methods
METHOTREXATE  Enzyme-mediated (multiplied) immunologic technique
 Inhibits DNA synthesis in all cells (EMIT)
 Leucovorin rescue: reverse action of methotrexate  Fluorescence polarization immunoassay (FPIA)
BUSULFAN CHROMATOGRAPHIC METHODS
 Treatment of leukemias and lymphomas prior to  best specimen: ____________________________
bone marrow transplantation 
ANTI-INFLAMMATORY/ANALGESICS  quantitatively identifies drugs by means of their Rf
values
 Commonly used analgesic, anti-pyretic and anti-  extraction of drugs is pH dependent – acidic drugs at
inflammatory drug pH 4.5 and alkaline drugs at pH 9.0
 Has anticoagulant property by inhibiting _____________ 
 Acute aspirin intoxication: common cause of fatal drug  ideal for separation of tricyclic antidepressants and
poisoning in children its metabolites
 Method: Trinder assay, HPLC, EMIT, Enzymatic assay  ____________________________________________ –
gold standard
 Commonly used analgesic and anti-pyretic drug

 Over dosage leads to hepatotoxicity TOXICOLOGY
 Method: HPLC ALCOHOLS
ETHANOL (GRAIN ALCOHOL)
 Lower risk of toxicities than salicylates and  most common abused drug
acetaminophen  antidote for chronic intoxication: diazepam (for
NEUROLEPTICS (ANTI-PSYCHOTIC MAJOR alcoholic mania)
TRANQUILIZERS)  specimen: serum (capillary and arterial blood are
preferred)
 Block the action of dopamine and serotonin in the limbic
system  methods for testing: enzymatic, gas-liquid
chromatography and electrochemical oxidation
 Used in the treatment of acute schizophrenia
 preferred method: enzymatic using alcohol
 2 classes: phenothiazines (chlorpromazine) and
dehydrogenase reagent
butyrophenones (haloperidol)
 Examples: risoperdal, olanzapine, quetiapine,  ____________________________ = symptoms of
aripiprazole alcohol intoxication begin to manifest
 ___________________________ = legally intoxicated
METHODOLOGIES FOR TDM
SPECIMEN CONSIDERATIONS  ___________________________ = presumptive
evidence of driving under influence of alcohol
 Specimen of choice: serum or plasma
Table 46. Stages of Ethanol Impairment
 Whole blood EDTA sample is required for cyclosporine
Alcohol (%, w/v) SxS
and
0.01 – 0.05 No obvious impairment, some changes
 tacrolimus test
observable on performance testing
 Timing of the specimen collection is the single most
0.03 – 0.12 Mild euphoria, decreased inhibitions,
important factor in TDM
some impairment of motor skills
 Trough concentrations are drawn immediately or 30
minutes before the next dose; lowest level of drug in
blood

Clinical Chemistry
0.09 – 0.25 Decreased inhibitions, loss of critical MERCURY

judgment, memory impairment,  inhibits catecholamine-0-methytransferase
diminished reaction time  has the ability to “amalgamate” – mix or merge with
0.18 – 0.30 Mental confusion, dizziness, strongly other substances
impaired motor skills (staggering,  forms of mercury: elemental/metallic mercury,
slurred speech) mercurous, mercuric and alkyl mercury
0.27 – 0.40 Unable to stand or walk, vomiting,  major toxic effect of elemental mercury: ____________
impaired consciousness ______________________________________________
0.35 – 0.50 Coma and possible death  major toxic effect of alkyl mercury: congenital Minimata
METHANOL (WOOD ALCOHOL) disease
 samples: whole blood and 24-hour urine
 commonly used solvent and a contaminant of
 method: ________________________
homemade liquors
LEAD
 Symptoms of intoxication: frank blindness (ocular
 blocks ________________________________________
toxicity) and metabolic acidosis
______________________________________________
 screening test: computation of osmolal gap
______________________________________________
 preferred method: GC-MS
 source: paints and gasoline
ISOPROPANOL (RUBBING ALCOHOL)
 indications of toxicity: urinary aminolevulinic acid, free
 metabolized by hepatic alcohol dehydrogenase into RBC proporphyrin and presence of
acetone ________________________________________ in RBC
 preferred method: gas chromatography  lead chelators: EDTA and dimercaptosuccinic acid (DMA)
 antidote: activated charcoal  characteristic “wrist drop” or “foot drop” manifestation
ETHYLENE GLYCOL (1,2 – ETHANEDIOL) (peripheral neuropathy)
 common constituent of hydraulic fluid and anti-freeze  samples: whole blood (quantitative testing), urine
(recent lead exposure), hair
 converted to oxalic acid and glycolic acid by hepatic
alcohol dehydrogenase LABORATORY TESTS FOR LEAD
 indication of toxicity: deposition of calcium oxalate 1. Screening test - Zinc protoporphyrin test
crystals in renal tubules (Fluorometric) δ-ALA dehydratase test (sensitive)
 major metabolite: glycolic acid (cause of acute toxicity 2. In-vivo x-ray fluorescence of bones
and death) 3. Atomic absorption spectrophotometry
 preferred method: HPLC 4. Inductively couples plasma emission spectrophotometry
SPECIMEN PRECAUTION 5. Anodic stripping voltammetry
DRUGS OF ABUSE
 Specimen must be capped at all times to avoid alcohol
 Designer drugs – are modified forms of established drugs
evaporation
of abuse
 prior to collection, alcohol-free skin cleanser must be
used instead of isopropanol
CARBON MONOXIDE  therapeutically used for the treatment of narcolepsy and
 is a colorless, odorless, tasteless gas; very toxic substance attentional deficit disorder
 binds with heme proteins: cytochromes, hemoglobin and  cause the release of dopamine from brain leading to
myoglobin “pleasant feeling” or “high” among users
 higher affinity for hemoglobin that does oxygen (200x  3,4-methylenedioxymethamphetamine (MDMA or
faster than oxygen) ‘ecstasy’) – a derivative of methamphetamine, is a
 indication of acute toxicity: “________” color of the face popular recreational abused drug
 sample for testing: EDTA whole blood  cause of false positive reaction: presence of
 definitive method: ___________________________ antihistamine
(carboxyhemoglobin measurement) ANNABOLIC STEROIDS
CYANIDE  chemically related to testosterone; improves athletic
 super toxic substance (fast acting toxin) and death may performance by increasing muscle mass
occur in less than an hour CANNABINOIDS
 inhibits cellular respiration, electron transport and ATP
formation  naturally-occurring cannabinoids: marijuana and
hashish
 indication of toxicity: “___________________________”
breath and altered mental status  __________________________________ : most potent
component or the psychoactive substance of marijuana;
 antidote: sodium thiosulfate, amyl and sodium
lipophilic

Clinical Chemistry
 THC-COOH can be detected in urine for 3-5 days; up to 4

weeks for chronic users METHODS FOR IDENTIFYING AND MEASURING
 urinary metabolite: 11-nor-deltatetrahydrocannabinol ABUSED DRUGS
(THC-COOH) 1. Enzymatic – alcohol is measured from blood using
COCAINE (CRACK) alcohol dehydrogenase as reagent
 used as _______________________________________ 2. Capillary electrophoresis – different analyte selectivity is
 derived from coca plant (Erythroxylon) and used as based on different physicochemical principles of
additive to some foods separation without changes in instrumental hardware.
 inhibitor: Prozac 3. Homogenous immunoassay – assay is done in one

solution without separation
 treatment for cocaine addiction: Benzodiazepine
 Enzyme mediated immunologic technique (EMIT) –
 urine metabolite: ______________________________
uses enzyme-labeled drug that competes with the
 not considered as an addictive drug – does not reflect
drug in sample
true dependence
4. Chromatographic methods
 easily passes the placenta and mammary glands resulting
 Thin layer chromatography – uses serum, urine or
to mental retardation, slow mental development and
gastric fluid for analysis
drug dependence in newborns
 Liquid chromatography-Mass spectroscopy (LC-MS)
 can cause sudden death due to direct toxicity in
– used to confirm positive test results from a
myocardium (cardiac toxicity)
screening assay
OPIATES
 High performance liquid chromatography (HPLC) –
 capable of analgesia, sedation and anesthesia used as an alternative to GC-MS in definitive
 derived chemically from opium poppy identification of drugs
 naturally occurring opiates: opium, morphine and  Gas chromatography
codeine o Gas liquid chromatography (GLC) – legally
 chemically modified opiates: heroine, hydormorphone accepted method for ethanol testing
and oxycodone o GC with infrared spectroscopy – detection of
 commonly tested opiates: _______________________ amphetamines
 major metabolites of heroine: N-acetylmorphine o Gas chromatography-Mass spectroscopy – gold
(heroin) and morphine standard for confirmation of screening tests;
 antagonist for opiate overdose: naloxone (narcan) allows for detection of low levels of drugs
 Codeine is ________________________________.
 Morphine is a metabolite of heroin used in the treatment REFERENCES
of congestive heart failure.  Rodriguez, Maria Teresa. Clinical Chemistry Handbook for
 Heroin is highly addictive (true physical dependence) medical technologists. Cattleya Star Copy Center and
PHENCYCLIDINE Book Binding, 2014
 Coderes, Errol. March 2018 Medical Technology Board
 physiologic effects: analgesia and anesthesia
Exam Final Coaching Notes. Pioneer Educational Review
 major metabolite: phencyclidine HCl
Center, Manila, 2018
 mode of treatment: isolation (kept in quiet, dark room)
 Policarpio, Judea Marie. March 2018 Medical Technology
SEDATIVE HYPNOTICS
Board Exam Final Coaching Notes. ACTS Review Center,
 examples: barbiturates and benzodiazepines Manila, 2018
 Commonly abused barbiturates: secobarbital,  Plaza, Xenia. March 2019 Medical Technology Board
pentobarbital, phenobarbital and thiopental Exam Clinical Chemstry Review Notes. Remedios Trinidad
 commonly abused benzodiazepines: diazepam (valium), Romualdez Medical Foundation, 2019
chlordiazepoxide (Librium) and lorezepam (Ativan)
 major metabolite: secobarbital (barbiturates)
 barbiturate chemoadsorbent: activated charcoal
LYSERGIC ACID DIETHYLAMIDE (LSD, LYSERGIDE)
 one of the most potent pharmacologic materials known

 most common adverse reaction: panic reaction - “bad
trip”
 toxic effects: blurred/ “undulating” vision and
synesthesia

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CLINICAL CHEMISTRY MTAP

BY: GEORGE VINCENT GELLENA, RMT

CONTENTS: Table 1. – Types of Error

COLLEGE OF MEDICAL LABORATORY SCIENCE | 1 | 34

7. __________________ – Displays output of the detection

Westgard Errors on LJ Chart

 Measurement of light transmitted by a solution to

COLLEGE OF MEDICAL LABORATORY SCIENCE | 2 | 34

 Measures light absorbed by atoms dissociated by heat; GAS CHROMATOGRAPHY

 Determines the amount of light emitted by a molecule

Optical Arrangements of Nephelometry and

 Migration of charged particles in an electric field

 Measures absorbance of stain  Chemical reaction yields electronically excited

 Separation of soluble components based on physical

COLLEGE OF MEDICAL LABORATORY SCIENCE | 3 | 34

ELECTROCHEMISTRY TECHNIQUES CARBOHYDRATES

COLLEGE OF MEDICAL LABORATORY SCIENCE | 4 | 34

 Glycemic factors such as glucagon are released when

 Severe DM, the ratio of β-hydroxybutyrate to  Diagnostic Criteria for GDM

COLLEGE OF MEDICAL LABORATORY SCIENCE | 5 | 34

GLUCOSE METHODOLOGIES  Guidelines for OGTT

 Short term glucose control (3 – 6 weeks)

COLLEGE OF MEDICAL LABORATORY SCIENCE | 6 | 34

VALUES TO REMEMBER o Catalyzes the esterification of cholesterol (HDL)

COLLEGE OF MEDICAL LABORATORY SCIENCE | 7 | 34

MINOR LIPOPROTEINS ENZYMATIC METHODS

COLLEGE OF MEDICAL LABORATORY SCIENCE | 8 | 34

IIa (Familial High LDL (High Cholesterol) FORMULA FOR LDL-C

COLLEGE OF MEDICAL LABORATORY SCIENCE | 9 | 34

 Friedewald Method (Indirect)

 TERMS:  transports iron to its storage sites

 binds heme released by degradation of hemoglobin;

COLLEGE OF MEDICAL LABORATORY SCIENCE | 10 | 34

 alternative test to chemical analysis of serum proteins

COLLEGE OF MEDICAL LABORATORY SCIENCE | 11 | 34

SERUM PROTEIN ELECTROPHORESIS  when the substrate concentration reaches a

COLLEGE OF MEDICAL LABORATORY SCIENCE | 12 | 34

Table 14. Classification of Enzymes

1. Emil Fisher’s (Lock and Key Theory) – the shape of the

COLLEGE OF MEDICAL LABORATORY SCIENCE | 13 | 34

METHODOLOGIES Table 16. Summary of ACP Methods

COLLEGE OF MEDICAL LABORATORY SCIENCE | 14 | 34

 ALT is slightly increased in obstructive jaundice but METHODOLOGIES

COLLEGE OF MEDICAL LABORATORY SCIENCE | 15 | 34

COLLEGE OF MEDICAL LABORATORY SCIENCE | 16 | 34

INDIRECT METHOD  Normal BUN to Creatinine Ration = _________________

COLLEGE OF MEDICAL LABORATORY SCIENCE | 17 | 34

 used in the diagnosis of hepatic failure (hepatic coma)

 Reference Range: 19 – 60 ug/dL

1. When RBCs are destroyed in the spleen, hemoglobin is

COLLEGE OF MEDICAL LABORATORY SCIENCE | 18 | 34

5. Protoporphyrin is broken down into biliverdin, and then  Diazo Reagents

COLLEGE OF MEDICAL LABORATORY SCIENCE | 19 | 34

3. _________________________________ – Bilirubin  Diarrhea, Excess fluid loss

 otherwise known as “kalium”

 Aldosterone deficiency, Ketonuria  Nephritis, Renal tubular acidosis,

COLLEGE OF MEDICAL LABORATORY SCIENCE | 20 | 34

COLLEGE OF MEDICAL LABORATORY SCIENCE | 21 | 34

 Mutations of Vitamin D receptor MAGNESIUM

 ______________________ related to calcium; maximally FACTORS AFFECTING MAGNESIUM LEVELS IN BLOOD

absorbed in the _________________________  __________________________________ - increases

  Acute or Chronic Renal Failure

COLLEGE OF MEDICAL LABORATORY SCIENCE | 22 | 34

absorbed in the _  __________ - increases