Clinical Chemistry Board Review

Photometric Detection Methods

1. Spectrometry-Chemical reaction produces a colored product that absorbs light of a

certain wavelength. Light absorbance is proportional to the concentration of the analyte

2. Atomic absorption (AA)-Measures light absorbed by ground state atoms

3. Fluorometry-Atoms absorb light of a specific wavelength and emit light of a longer

wavelength (lower energy)

4. Chemiluminescence-Clinical reaction that produces light via oxidation of luminal,

acridinium esters, or dioxetanes

5. Turbidimetry-Measure reduction in light transmission by particles in suspension’

6. Nephelometry-Similar to turbidity; light is measured at an angle from the light source

Chromatography Methods

1. Thin layer chromatography-solvent coated glass or plastic plate, closed solvent

chamber, organic solvent

2. High performance liquid chromatography (HPLC)-Instrument has a solvent pump,

injection port for sample, separation column, detector, integrator, and recorder

3. Gas chromatography (GC)-Gas, injection port, separation column, oven, detector,

recorder. Sample must be vaporized when injected into instrument, unlike HPLC

Other techniques:

1. Ion selective electrode (ISE)

-Potential difference between 2 electrodes is directly related to concentration of analyte

2. Osmomitry
-Determine osmolarity (measurement of dissolved particles in solution
-Based on freezing point depression in most clinical labs, also vapor pressure
instruments available

3. Electrophoresis
-Separation of charged particles in an electrical field by molecular weight or isoelectric
point
Steps in Automated Analysis

a. Sample ID-Bar coded blood collection tubes

b. Test Selection-What assay (s) is/are to be performed on the sample?

c. Sampling-Usually closed tube sampling directly from blood collection tubes

-Vitros® system uses dry slides, no wet reagents

d. Reagent delivery-Usually by syringes, pumps, pressurized reagent containers

e. Chemical reaction-Mixing and incubation

f. Measurements-UV and visible spectroscopy, ISEs, fluorescence polarization,

chemiluminescence, bioluminescence

g. Data Handling-Concentration of analyte derived from calibration curve stored in the

analyzer

h. Reporting of Results-Interface between lab information system (LIS) and hospital

computer system

i. Troubleshooting-Can be done remotely in the modern analyzers

Commonly Ordered Chemistry Panels

1. Basic Metabolic Panel (BMP): Na+, K+, Cl-, CO2, glucose, creatinine, BUN, Ca2+

2. Comprehensive Metabolic Panel: All of the above analytes + albumin, total protein,
ALP, AST, and bilirubin

3. Electrolyte Panel: Na+, K+, Cl-, CO2

4. Hepatic Function Panel: Albumin, AST, ALT, ALP, total and direct bilirubin, total
protein

5. Lipid Panel: Total cholesterol, HDL, LDL, triglycerides

6. Renal Function Panel: Na+, K+, CO2, glucose, BUN, creatinine, Ca2+, albumin,
phosphate
Carbohydrates, Lipids and Proteins

1. Glucose (fasting) 70-99 mg/dL

-↑in diabetes mellitus, endocrine disorders, acute stress, pancreatitis
-↓ in insulinoma, insulin induced hypoglycemia, hypopituitarism

2. Total cholesterol <200 mg/dL

-Predictive for coronary artery disease (CAD) in conjunction with HDL and LDL

3. HDL >60 mg/dL

-Inversely related to CAD risk

4. LDL <100 mg/dL

-Risk factor associated with CAD

5. Triglycerides <150 mg/dL

-Elevated levels indicate risk factor for CAD

6. Total protein 6.4-8.3 g/dL

-↑ in dehydration, chronic inflammation, multiple myeloma
-↓ in nephritic syndrome, malabsorption, overhydration, hepatic insufficiency,
malnutrition, agammaglobulinemia

7. Albumin 3.5-5.0 g/dL

-↑ in dehydration
-↓in malnutrition, liver disease, nephritic syndrome, chronic inflammation

8. Urine microalbumin 50-200 mg/24hrs

-↑ in diabetics at risk of nephropathy
-↓ predictive of diabetic nephropathy

Tests for diabetes mellitus (DM)

1. Random plasma glucose >200 mg/dL is diagnostic of DM

2. Fasting plasma glucose >126 on 3 occasions (patient fasts for 8 hours)

3. 2-hour plasma glucose: 75 g of glucose is given initially to patient >200 mg/dL

4. Oral glucose tolerance test (OGTT): 75 g glucose load given to patient Results:
Fasting level >92 mg/dL, 1 hour time >180 mg/dL, 2 hour time >153 mg/dL

5. Hemoglobin A1C: No patient prep or fasting is needed>6.5% is diagnostic of

diabetes mellitus
Increased Lab Values Observed in Uncontrolled Diabetes Mellitus

a. Blood glucose
b. Urine glucose
c. Glycohemoglobin
d. Ketones (in blood and urine)
e. Anion gap
f. BUN
g. Serum and urine osmolarity
h. Cholesterol
i. Triglycerides

Decreased Lab Values in DM

a. Bicarbonate
b. Blood pH

Metabolic Syndrome-Risk factors promoting development of atherosclerotic disease and

Type 2 diabetes mellitus:
↓ HDL-C
↑ LDL-C
↓ Triglycerides
↑ Blood glucose
↑ Blood pressure

Aminoacidopathies

1. Phenylketoneuria (PKU): Deficiency of enzyme that converts phenylalanine to

tyrosinephenylpyruvate appears in blood and urine

2. Tyrosinemia: Tyrosine metabolic disorder where tyrosine and its metabolites are
excreted in the urine

3. Alkaptonuria: Tyrosine, phenylalanine disorder that causes a buildup of homogentisic

acid

4. Maple syrup urine disease (MSUD): Enzyme deficiency leading to buildup of leucine,
isoleucine and valine

5. Homocystinuria: Deficiency in enzyme that metabolizes methionine

-Methionine and homocysteine builds up in plasma and urine

6. Cystinuria: Defect in renal reabsorption of cystine

-Causes recurring bladder stones
Protein Electrophoresis (In order of separation):

Albumin, α1, α2, β, γ (albumin is the largest peak)

Acute inflammation: ↑α1, α2

Chronic infection: ↑α1, α2, γ
Cirrhosis: Beta-gamma bridging
Monoclonal gammopathy: ↑ in one immunoglobulin (M spike), ↓in other fractions
Nephrotic syndrome: ↓albumin, ↑α2
Alpha-1-antitrypsin: ↓α1

Non-Protein Nitrogen Compounds

a. BUN: 8-26 mg/dL ↑ in kidney disease, ↓ in liver disease and over-hydration

b. Creatinine: 0.7-1.5 mg/dL ↑ in kidney disease

c. Uric acid: M: 3.5-7.2 mg/dL; F: 2.6-6.0 mg/dL ↑ in gout, kidney failure, leukemia,
lymphoma, ketoacidosis, lactate excess; ↓ in ACTH administration

d. Ammonia: 19-60 mg/dL ↑ in liver disease, hepatic coma, renal failure, Reye’s
syndrome

Electrolytes

a. Sodium (Na+) 136-145 mmol/L

↑ due to increased intake, hyperaldosteronism, burns, diabetes insipidus
↓ due to renal or extrarenal loss (vomiting, diarrhea), increased extracellular fluid
volume

b. Potassium (K+) 3.5-5.0 mmol/L

↑ due to increased intake or decreased excretion, crush injuries, metabolic acidosis
-Increased K+ can cause cardiac arrhythmia, cardiac arrest
↓ due to increased GI and urinary losses; diuretics, metabolic alkalosis
-Decreased K+ can cause muscle weakness, paralysis, cardiac arrhythmia, death

c. Chloride (Cl-) 98-107 mmol/L

↑ due to same conditions as ↑ Na+, excess loss of HCO3-
↓ due to prolonged vomiting, diabetic ketoacidosis, respiratory acidosis (compensated)

d. Total CO2 23-29 mmol/L

↑ in metabolic alkalosis, respiratory acidosis (compensated)
↓ in metabolic acidosis, respiratory alkalosis (compensated)
e. Magnesium (Mg2+) 1.6-2.6 mg/dL
↑ in renal failure, ↑ intake (antacids), dehydration, bone cancer, endocrine disorders
-Increased can cause respiratory arrest, coma, cardiac abnormalities, paralysis
↓ in GI disorders, severe illness, renal losses, endocrine disorders
-Decreased can cause tremors, tetany, paralysis, psychosis, coma, cardiac arrhythmias

f. Calcium (Ca2+) 8.6-10.0 mg/dL (total); 4.6-5.1 mg/dL (ionized)

↑ in primary hyperparathyroidism, cancer, multiple myeloma
-Increased causes weakness, coma, GI symptoms, renal calculi
↓ in hypothyroidism, malabsorption, vitamin D deficiency, renal tubular acidosis
-Decreased causes tetany, seizures, cardiac arrhythmias

g. Phosphorus 2.5-4.5 mg/dL

↑ in renal disease, hypoparathyroidism
↓ in hyperparathyroidism, vitamin D deficiency, renal tubular acidosis

h. Lactate (lactic acid) 4.5-19.8 mg/dL

-Sign of decreased O2 delivery to the tissues

Iron Studies

1. Iron M: 65-175 μg/dL F: 50-170 μg/dL

↑ in Iron overdose, hemochromatosis, sideroblastic anemia, hemolytic anemic, liver
disease
↓ Iron deficiency anemia (IDA)

2. Total iron binding capacity (TIBC) 250-425 μg/dL

↑ in IDA
↓ in iron overdose, hemochromatosis

3. % transferring saturation (% saturation) 20-50%

↑ Iron overdose, hemochromatosis, sideroblastic anemia
↓ in IDA

4. Transferrin 200-360 mg/dl

↑ in IDA
↓ in iron overdose, hemochromatosis, chronic infections, malignancies

5. Ferritin M: 20-250 μg/dL F: 10-120 μg/dL

↑ in iron overload, hemochromatosis, chronic infections, malignancies
↓ in IDA
Enzymes of Clinical Significance

1. Acid phosphatase (ACP)

↑ in prostate cancer

2. Alkaline phosphatase (ALP)

↑ in liver, bone disease, viral hepatitis (↑↑), acute myocardial infarction, muscular
dystrophy

3. Aspartate aminotransferase (AST)

↑ in liver disease (hepatitis), AMI, muscular dystrophy

4. Alanine aminotransferase (ALT)

↑ in liver disease; more specific than AST
-Also found in RBCs (hemolysis will affect test results)

5. Gammaglutamyl transferase (GGT): Found in liver, kidneys, pancreas

↑ in all hepatobiliary disorders, chronic alcoholism

6. Lactate dehydrogenase (LDH): Found in liver, heart, skeletal muscle, RBCs

↑ in acute myocardial infarct, muscular dystrophy

7. Amylase (AMY): Found in pancreas, salivary glands

↑ in acute pancreatitis, abdominal diseases, mumps

8. Lipase
↑ in acute pancreatitis; levels parallel amylase, but may be elevated longer; more
specific than amylase for pancreatic disease

9. Glucose-6-phosphate dehydrogenase (G6PD): Found in RBCs

-Inherited deficiency can lead to drug induced hemolytic anemia

Enzymes Used in Diagnosis by Group

a. CK-MB-Cardiac disorders
b. AST, ALT, LDH-Hepatocellular disorders
c. ALP, GGT-Biliary tract disorders

Cardiac Markers: Duration of Elevation, Sensitivity, Specificity

1. CK-MB- ↑ 4-6 hours after MI symptoms (chest pain, etc.)

-Remains elevated 2-3 days
-Not entirely specific for MI

2. Cardiac troponin- ↑ 4-10 hours after MI symptoms

-Remains elevated for 4-10 days
-Very specific and sensitive for MI
3. Myoglobin- ↑ 1-4 hours after MI symptoms
-Remains elevated for 18-24 hours
-Sensitive but NOT specific!

Other Cardiac Tests

1. B-type natriuretic peptide (BNP)-Used to diagnose CHF

2. Cardiac C-reactive protein (CCRP)-High sensitivity CRP used to diagnose the risk of
cardiac disease

3. Total cholesterol <200 mg/dL

4. HDL >60 mg/dL

5. LDL <100 mg/dL

6. Triglycerides <150 mg/dL

Bilirubin and Hepatic Disease

1. Total bilirubin 0.2-1.0 mg/dL

↑ in liver disease, HDN, hemolysis
-Total bilirubin >20 mg/dL is associated with infant brain damage

2. Conjugated (direct) bilirubin <0.2 mg/dL

↑ in liver disease, obstructive jaundice
-Direct bilirubin is a glucuronide conjugate of bilirubin (H2O soluble)
Jendrassik-Grof method is used to assay total and conjugated bilirubin

-Indirect (unconjugated) bilirubin <0.8 mg/dL

-↑ in prehepatic, posthepatic and some types of hepatic jaundice

Unconjugated vs. conjugated bilirubin

1. Unconjugated bilirubin: Bound to protein, nonpolar, high affinity for brain tissue, not
H2O soluble
2. Conjugated bilirubin: Mono- and di-glucuronide forms + delta bilirubin
-Not bound to protein polar, present in urine
Pre-hepatic jaundice Hepatic jaundice Post-hepatic jaundice

↑ total bilirubin ↑ total bilirubin ↑ total bilirubin

N direct bilirubin Variable direct bilirubin ↑ direct bilirubin
N urine bilirubin Variable urine bilirubin Pos urine bilirubin
↑urine urobilinogen ↓ urine urobilinogen ↓ urine urobilinogen

Hormones
1. ACTH: (pituitary), stimulates production of adrenocortical hormones by the adrenal
cortex
↑ Cushing’s disease, diurnal variation (highest levels obtained in the AM)

2. Follicle stimulating hormone (FSH): (pituitary), production of eggs and sperm

-Regulated by hypothalamic GnRH

3. Luteinizing hormone (LH): (pituitary), maturation of the follicles and ova, production of
estrogen, progesterone, testosterone
↑ LH levels-increase before ovulation (sharp ↑ just before ovulation)

4. Growth hormone (GH): Stimulates protein synthesis, cell growth and cell division

5. Prolactin: Regulates lactation; regulated by hypothalamic PRF and PIF

6. TSH: Stimulates production of T3 and T4 by thyroid

↑ in hypothyroidism
↓ in hyperthyroidism

7. ADH: Posterior pituitary hormone-Release stimulated by increased osmolality

-Decreases blood volume and blood pressure
↓ in diabetes insipidus

8. Oxytocin: Posterior pituitary hormone-Causes uterine contractions during childbirth,

aids in lactation
-Not clinically useful to assay

9. Thyroid hormones: T4=thyroxine; T3=triiodothyronine

-Metabolic growth hormone regulated by TSH and bound to TBG
↑ in hyperthyroidism
↓ in hypothyroidism
Thyroid Function Testing Results

Primary hypothyroidism TSH↑ FT4↓ FT3↓

Secondary hypothyroidism TSH↓ FT4↓ FT3↓

Hyperthyroidism TSH↓ FT4↑ FT3↑

T3 thyrotoxicosis TSH↓ FT4 N* FT3↑

*N=normal

10. Calcitonin: Produced by the C cells of the thyroid-Inhibits reabsorption of Ca2+

-Used in the diagnosis of thyroid cancer

11. Parathyroid Gland

-Parathyroid hormone (PTH)-Regulates Ca2+ and phosphate
-Primary hyperthyroidism=↑PTH, ↑Ca2+, ↓phosphate
-Hypoparathyroidism=↓PTH, ↓Ca2+, ↑phosphate

12. Adrenal Hormones

Adrenal cortical hormones:
a. Aldosterone: Regulates reabsorption of Na+ in renal tubules
↑ levels cause hypertension (Na+ and H2O retention)
↓ levels cause severe H2O and electrolyte abnormalities

b. Cortisol: Carbohydrate, fat, and protein metabolism; H2O and electrolyte balance;
suppress inflammatory and allergic reactions
↑ in Cushing’s disease
↓ in Addison’s disease

Adrenal medulla hormones:

Epinephrine and norepinephrine: Stimulate the sympathetic nervous system
↑Pheochromocytoma (catecholamine producing tumor)
-Plasma and urine catecholamines, metanephrines, urine VMA

Reproductive Hormones
-Ovarian hormones:
a. Estrogens: Primary estrogen is estradiol (E2)-Regulates menstrual cycle, pregnancy
b. Progesterone: Preparation of uterus for ovum implantation, maintenance of
pregnancy
-Used in fertility studies and to assess placental function

-Placental hormones:
a. Estriol-Used to monitor fetal growth and development
b. Progesterone- Used to monitor fetal growth and development
c. HCG-Used to detect pregnancy, gestational trophoblastic disease, HCG producing
tumors
-Testicular hormones:
a. Testosterone-Male reproductive development

Pancreatic hormones

1. Insulin: Carbohydrate metabolism-causes increased movement of glucose into cells

for metabolism (decreases plasma glucose levels)
↓ in diabetes mellitus
↑ in hypoglycemia, insulinoma

2. Glucagon: Glycogenolysis, gluconeogenesis, lipolysis

↑ in glucagonoma
↑ in D.M., pancreatitis, trauma

Therapeutic Drugs

1. Analgesics (pain management): Salicylates, acetaminophen

2, Antiepileptics: Phenobarbital, phenytoin, valproic acid, carbamezapine, ethosuximide,

felbamate, galsapertin, lanotrigine

3. Antineoplastics: Methotrexate

4. Antibiotics: Aminoglycosides (amikacin, gentamycin, kanamycin, tobramycin,

vancomycin)

5. Cardioactives: Digoxin, disopyramide, procainamide, quinidine

6. Psychoactives: Tricyclic antidepressants, lithium

7. Immunosuppressants: Cyclosporine, tacrolimis (FK-506)

Toxic Drugs/Substances

1. Ethanol: Assay by GC, enzymatic methods

2. Carbon monoxide (CO): Assay by cooximeter, GC

3. Arsenic: Assay by atomic absorption (AA)

4. Lead: Assay by AA

5. Pesticides-Measurement of serum pseudocholinesterase

Drugs of Abuse Screen

-Amphetamines, barbiturates, benzodiazapines (valium), cannabinoids, cocaine,

methadone, opiates, phencyclidine, tricyclic antidepressants

-Method of detection: Usually immunoassays, confirm the results by using a different

assay technique

Common Tumor Markers

1. α-fetoprotein (AFP)
-Liver cancer, produced by fetal liver and re-expressed in some tumors; also increased
in hepatitis and pregnancy

2. Cancer antigen15-3 (CA 15-3) + cancer antigen 27,29 (CA 27,29)

-Breast cancer marker; can be increased in other cancers

3. Cancer antigen 19-9 (CA19-9)

-Pancreatic cancer marker, can be increased in other cancers and non-cancerous
conditions

4. Cancer antigen 125 (CA 125)

-Ovarian cancer marker, can also be elevated in other cancers

5. Carcinoembryonic antigen (CEA)

-Colorectal cancer marker; fetal antigen re-expressed in tumors, can be increased in
other cancers, non cancerous conditions and smokers

6. Human chorionic gonadotropin (HCG)

Ovarian and testicular marker, also gestational trophoblastic disease
↑ in pregnancy (used in clinical lab to determine pregnancy)

7. Prostate specific antigen (PSA)

-Prostate cancer marker; some patients with prostate cancer don’t have elevated PSA
-PSA can be elevated in other conditions
-Measure free PSA if PSA is borderline

8. Thyroglobulin
-Thyroid cancer marker
-Also elevated in other diseases; antithyroglobulin antibodies should be measured at the
same time
Acid-Base Homeostasis

1. Acidosis (acidemia)
Blood pH <7.3
↓ HCO3-:H2CO3 ratio (20:1 is normal)
-May be due to ↓ in HCO3- (metabolic acidosis) or ↑ in H2CO3 (respiratory acidosis)

2. Alkalosis (alkalemia)
Blood pH>7.42
↑ HCO3- : H2CO3 ratio
-May be due to ↑ in HCO3- (metabolic alkalosis) or ↓ in H2CO3 (respiratory alkalosis)

-Compensated acidosis or alkalosis

Kidneys compensate for respiratory problem; lungs compensate for metabolic problem

Acid-Base Imbalances

Respiratory acidosis pH↓ pCO2↑ HCO3- N*

Respiratory alkalosis pH↑ pCO2↓ HCO3- N
Metabolic acidosis pH↓ pCO2↓ HCO3- ↓
Metabolic alkalosis pH↑ pCO2↑ HCO3- ↑

*N=normal

Blood Gas Normal Values

a. Blood pH=7.35-7.45
b. pCO2=35-45 mmHg
c. PO2=80-100 mmHg
d. HCO3- =22-26 mmol/L
e. Base excess= -2 to +2 Difference between titratable bicarbonate of patient’s sample
and a normal blood sample
f. O2 saturation=94-100% (Amount of oxygenated hemoglobin)

Common Blood Gas Errors

1. Specimen exposed to air ↓pCO2 ↑O2 ↑pH

2. Specimen held at room temperature for >30 min. ↓pCO2 ↓pH ↑pCO2
Clinical Chemistry Formulas

1. A/G Ratio= Albumin/(total protein-albumin) Normal range=1-2.5

2. Amylase: creatinine clearance ratio= urine amylase (U/L)x serum creatinine (mg/L) ÷
serum amylase (U/L) x urine creatinine (mg/L)

3. Anion gap= Na+ - (Cl- +HCO3-) Normal=7-16

or AG= (Na++K+) - (Cl- +HCO3-) Normal=10-20

4. BUN/Creatinine ratio = BUN/creatinine Normal=10-20

5.Creatinine clearance= urine creatinine(mg/dL) x urine ml per 24 hr/1440 ÷ plasma

creatinine (mg/dL) x 1.73 ÷ body surface area M: 97-137 ml/min; F: 88-128ml/mi n

6. Indirect (unconjugated) bilirubin=total bilirubin- direct (unconjugated) bilirubin N=<0.2

mg/dL

Lab Calculations

1. mEq/L= (mg/dL ÷ GEW) x 10 GEW= atomic weight ÷ valence of element (i.e. Ca2+)

2. mmol/L= (mg/dL ÷ GMW) x 10

3. mmol/L= mEq/L ÷ valence

4. Molarity (M) = grams per liter ÷ GMW

5. % Concentration = grams or ml per 100 ml

6. V1C1 = V2C2