Professional Documents
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BB Notes
BB Notes
BB Notes
1492 Pope Innocent VIII – recipient of first blood transfusion in history, but he & the donors died.
1869 Braxton Hicks – recommended sodium phosphate as nontoxic anticoagulant
1901 Karl Landsteiner – discovered ABO blood groups and explained transfusion reaction.
1902 Von Decastello and Sturli – discovered the fourth ABO blood group, AB
1913 Edward E. Lindemann – used vein-to-vein transfusion of blood by using multiple syringes
1913 Lester Unger – designed syringe-valve apparatus; permitted unassisted physician
1914 Albert Hustin – reported the use of sodium citrate Anticoagulant
1915 Richard Lewisohn – determined the minimum amount of citrate anticoagulant to be used
1916 Rous and Turner – introduced citrate-dextrose solution for preservation of blood
1941 Charls Drew – his techniques has led to widespread system of blood banks in WWII
1943 Loutit and Mollison – introduced the formula for acid-citrate-dextrose (ACD)
1957 Gibson – introduced an improved preservative solution, citrate-phosphate-dextrose (CPD)
1 unit of whole blood = 450 mL of blood (one pint) and 63 mL of Ac-preservative solution
Storage
Name Abbreviation Shelf-life temperature
Acid-citrate-dextrose ACD 21 1 – 6 °C
Citrate-phosphate-dextrose CPD 21 1 – 6 °C
Citrate-phosphate-adenine CPDA-1 35 1 – 6 °C
Citrate-phosphate-double dextrose CP2D 21 1 – 6 °C
Additive Solutions
Prolongs red cell survival for up to 42 days
Generally contains: SALINE, ADENINE, GLUCOSE, MANNITOL (AS-1, AS-5)
100 mL additive solution is used for a 450 mL blood bag
Solution must be added to RBCs within 72 hours of storage
Rejuvenation Solutions
used to restore levels of ATP and 2,3-DPG levels in red cells stored in CPD, CPDA-1, AS-1
Generally contains: PHOSPHATE, INOSINE, PYRUVATE, ADENINE
Solution can be used up to 3 days after expiration of red cells
REJUVESOL : the FDA-approved rejuvenation solution in US
Red Cell Freezing
primarily used for autologous units and storage of rare blood types
involves addition of cryoprotective agent to red cells that are < 6 days old.
Glycerol is most commonly used as cryoprotective agent
Long term storage : 10 yrs. at -65 °C
Deglycerolization
to be done prior transfusion to avoid infusing the hypertonic glycerol
red cell washing with decreasing conc. of saline (12 % saline, 1.6 % saline, 0.2 % dextrose with
saline) or commercially available cell-washing system.
*24 hrs. = outdating period for thawed red cells stored at 1-6 °C
No agglutination 0
3. Effect of pH
Ideal pH 6.5 to 7.5
4. Effect of temperature
IgM react best at cold temperature (4-22°C)
IgG react best at warm temperature (37°C)
Zeta potential
expression of the difference in electrostatic potential at the surface of the red cells & the ionic
cloud of positive cations that are attracted to the negative charges on the surface
ID of IgM Antibody
1. Saline
4-22°C immediate spin (IS) up to 60 mins.
primarily identifies IgM
ID of IgG Antibody
1. Protein Media
increases the dielectric constant, w/c reduces the zeta potential
Examples of protein media:
o 22 % albumin
causes agglutination by adjusting zeta potential between Red cells
incubation at 37°C for 15 – 60 mins
o Polybrene
can detect ABO incompatibility & clinically significant IgG alloAb
o Polyethylene glycol
reduces false-positive reactions
more effective than albumin, LISS, or manual polybrene
incubation at 37°C for 10 – 30 mins; cell washing before IAT
NOTE: Test mixture CANNOT be centrifuged & examined for aggl’n
o Polyvinylpyrrolidone (PVP)
o Protamine
3. Enzymes
ENHANCES antibody reactivity to Rh, Kidd, P1, Lewis and I antigens
DESTROYS reactivity to red cell antigens Fya, Fyb, M, N, & S.
Examples:
o Ficin (plant)
o Papain (papaya)
o Trypsin (pig stomach)
o Bromelin (pineapple)
DTT & 2 ME
sulfhydryl compounds that break the disulfide the bonds of the J chain of IgM molecule,
but leave IgG molecule intact
ZZAP reagent
thiol reagent + proteolytic enzyme
causes dissociation of IgG molecules from the surface of sensitized red cells
SEROLOGIC SYSTEMS USED IN TRADITIONAL LAB. METHODS
FOR RED CELL ANTIBODY DETECTION
ANTIGLOBULIN TEST IgG AHG has specificity for the Fc portion of heavy chain of IgG
It acts as a bridge cross-linking red cells.
ANTIGLOBULIN TEST
“Coombs Test”, discovered by Robin Coombs in 1945
based on the principle that anti-human globulins (AHG) obtained from immunized nonhuman
species bind to human globulins such as IgG or complement, either in free state or attached to
RBCs
Primarily detect IgG and / or complement-sensitized RBCs.
Polyspecific AHG
contains anti-IgG & anti-C3d component of complement system.
Monospecific AHG
contain only one Ab specificity: anti-IgG or anti-C3d
Polyclonal AHG
produced by immunizing a colony of rabbits (for large volume productions)
there is heterogeneity of IgG molecules (use of serum from many donors)
there is also pooling of anti-IgG from many immunized rabbits
Monoclonal AHG
technique devised by Kohler & Milstein
prepared by “Hybridoma Technology”; uses mice as lab animals
has been used to produce AHG with high-titer antibodies and well-defined specificities
Advantage: pure & uncontaminated reagent
4. Incubation Time
for cells suspended in saline: 30 – 120 mins.
for LISS: 10 – 15 mins. (extended incubation = decreased sensitivity)
5. Washing of Cell
wash RBCs 3 times before immediate addition of AHG
purpose is to REMOVE free unbound globulins
inadequate washing will result to false (-) results, because of neutralization by free globulins
washing should be performed in as short time as possible to minimize elution of low-affinity Abs
Karl Landsteiner
first described blood groups A, B, O (1901)
first individual to perform forward & reverse grouping
A B O AB
Antigen present on A antigen B antigen No A or B antigen A and B antigens
red cells
Antibody present Anti-B Anti-A Anti-A, Anti-B, NO Anti-A and
on serum Anti-A,B NO Anti-B
Type O
both phenotype & genotype are the same
individual would have to be HOMOZYGOUS for O gene.
ABO MATINGS
A A
Ex. Father = BO B AB AB Chances of Offspring:
Mother = AA AB = 50%
O AO AO AO = 50%
ABO GENETICS
ABO genes code for the production of SPECIFIC GLYCOSYLTRANSFERASES (enzymes)
that add sugars to a basic precursor substance
The ABH glycolipid antigens are built upon a common carbohydrate residue w/c represents a
PARAGLOBOSIDE (Precursor Substance)
Bombay Phenotype
LACKS normal expression of ABH antigen because α-2-L-fucosyltransferase is NOT produced
devoid of antigens of the ABO system
Immunodominant sugar
sugar that occupy the terminal positions of the precursor chain & confer the blood group specificity
GalNAC D-galactose
Immunodominant
Gene Glycosyltransferase (enzyme) Sugar donor sugar Antigen
α-3-N-acetylgalactosaminyl
A transferase UDP-GalNAc N-acetyl-galactosamine A
O allele
AMORPH
H substance remains unmodified
Blood Group O has the HIGHEST CONCENTRATION OF H ANTIGEN
ABH antigens
also referred to as “Histoblood Group Antigens”
present in all organs if the human body
SeSe or Sese
“Secretor” (ABH blood group-specific substances can be found in all body secretions)
78% of random population
sese
“Nonsecretor”
22% of random population
A Subgroups
A1
80% of all Group A individuals
very potent gene (810,000 – 1,170,000 antigen sites)
greater activity of GalNAC
A2
20% of all Group A individuals
produces only 240,000 – 290,000 antigen sites
lesser activity of GalNAC
A2 A Positive Negative
Sources of Anti-A1
Absorbed serum from Group B individuals
DOLICHOS BIFLORUS (Anti-A1 Lectin)
o agglutinates A1 and A1B
Lectins
seed extracts that agglutinate human cells w/ some degree of specificity
o DOLICHOS BIFLORUS (Anti-A1 Lectin)
o ULEX EUROPAEUS (anti-H Lectin)
Reactivity of Anti-H Antisera or Lectin w/ ABO Blood Groups:
* O = greatest amount of H Ag
* A1B = least amount of H Ag
BOMBAY PHENOTYPE
very rare, does NOT produce α-2-L-fucosyltransferase necessary for formation of H structure
can be genetically expressed as hh or Hnull or Oh
ABH antigens are absent, no agglutination w/ Anti-A, Anti-B or Anti-H
They are A,B,H nonsecretor (No ABH substance in saliva)
A Recessive mode of inheritance
Red cells of Bombay will NOT REACT w/ anti-H Lectin
Red cells of Bombay are compatible only w/ the serum from another Bombay individual
Group 1 Discrepancy
WEAKLY REACTING OR MISSING ANTIBODIES
Reason: Patient has depressed antibody production
Examples:
o Newborns
o Elderly patients
o Patients demonstrating Hypogammaglobulinemia caused by:
CLL
Malignant lymphoma
Immunosuppressive drugs
o Patients w/ Congenital Agammaglobulinemia
o Patients w/ Immunodeficiency states
o Patents w/ bone marrow transplantations
o CHIMERISM (presence of two cell populations in a single individual & occurs in twins)
Resolutions:
o Incubate px serum w/ reagent A1 & B cells at room temp. for 15 – 30 mins
Group 2 Discrepancy
WEAKLY REACTING OR MISSING ANTIGENS
Examples:
o Subgroups of A or Subgroups of B
o Leukemia – may yield weakened A or B antigen
o Hodgkin’s disease – mimic the depression of Ag found in leukemia
o Acquired B phenomenon
o Excess amounts of BGSS (blood-group specific-soluble substances) present in plasma
Resolution:
o Washing the patient’s cells free of the BGSS w/ saline should alleviate the problem
Group 3 Discrepancy
PROTEIN OR PLASMA ABNORMALITIES
Result in Rouleaux Formation
Examples:
o Elevated levels of globulin:
Multiple Myeloma
Waldenstrom’s Macroglobulinemia
Plasma cell dyscrasias
Advanced cases of Hodgkin’s lymphomas
o Elevated levels of fibrinogen
o Plasma expanders such as Dextran and Polyvinyl-pyrrolidone
o WHARTON’S JELLY
Resolution:
o Performing a saline dilution or saline replacement technique will free the cells in the
case of rouleaux formation in the reverse type
o In true agglutination, red cell clumping will still remain after addition of saline
Group 4 Discrepancy
MISCELLANEOUS PROBLEMS
Examples:
o Polyagglutination
o Cold Reactive antibodies (allo & auto)
o Warm autoantibodies
o Unexpected ABO isoagglutinins
o Ab other than Anti-A and Anti-B
o RBCs with cis-AB phenotype
Polyagglutination
Red cells agglutinating w/ all human sera
Genetic inheritance or bacterial infection
Example:
o T polyagglutination
exposure of a hidden RBC Ag (T Ag) occurs in patient w/ bacterial / viral infection
cis-AB phenotype
refers to the inheritance of both AB genes from one parent carried on one chromosome, and an O
gene inherited from the other parent.
Genotype: ABO
Usually the B antigen yields a weaker reaction w/ the anti-B
Rh Genetics
The genes that control the system are autosomal codominant located on the short arm of
chromosome 1.
The Rh Ag are inherited as CODOMINANT ALLELES
o Offspring inherit one Rh haplotype from each parent
Rh Biochemical structure
Nonglycosylated protein (no carbohydrate attached to the protein)
Rh antigens are transmembrane polypeptides & are integral part of RBC membrane
Rh TERMINOLOGIES
D Gene D
Production
Close Linkage C/c Gene C/c
Pathway
D 85%
(d or absence of D) 15%
C 70%
c 80%
E 30%
e 98%
Agglutinogen
RBC surface
WEAK D
Red cells carrying Weak D Ag have historically been referred to as having the Du type
1. Genetic Weak D
inheritance of D genes that code for a weakened expression of the D antigen
appears to be COMPLETE, but few in number
seen mostly frequently in Blacks
2. C Trans
“Position Effect” or “Gene interaction effect”
the allele carrying D is trans C or in the opposite haplotype to the allele carrying C
o e.g. Dce / dCe
Interference does not occur when C gene is inherited in the cis position to D e.g. DCe / dce
They can receive D-positive cells with no adverse effects
3. D Mosaic
one or more parts of the D Ag is MISSING
Anti-D made by D-mosaic individuals can cause HDN or transfusion reactions
Rh Antibodies
IgG; react at 37ºC or in AHG
exposure to less than 1 mL of Rh(+) red cells can stimulate Ab production in an Rh(-) person
IgG1 and IgG3 are of the greatest clinical significance because RES rapidly clears red cells
coated with them in the circulation
Rh antibodies DO NOT BIND COMPLEMENT
o Rh antigens are not situated on the red cell surface this closely. Red cell destruction resulting from
antibodies is primarily EXTRAVASCULAR
Rhmod
exhibit features similar to those with Rhnull
however, the clinical symptoms are usually less severe
Exalted D
DO NOT demonstrate Cc and / or Ee reactivity
unusually strong D expression
represented as D-- / D--
Enhanced by Enzymes:
Kidd
Rh
Lewis
I
P1
Destroyed by Enzymes:
Duffy
M,N,S,s
Shows Dosage:
Duffy
M,N,S,s
Kidd
Rh
BLOOD DONOR DEFERRAL
PERMANENT
HIV (+) / AIDS
HBsAg (+) , anti-HBc (+)
Hemophilia, von Willebrand’s disease, receiving clotting factors
Sickle cell anemia, Thalassemia, Polycythemia
Kaposi sarcoma
Malignant Tumors (except for basal cell carcinoma of skin & carcinoma in situ of cervix)
Chronic cardiopulmonary, liver or renal disease
History of Babesiosis or Chaga’s disease
Recipient of human dura mater grafts, pituitary-derived growth hormone
Donors who injected bovine insulin manufactured since 1980 from cattle in the UK
Have taken the drug TEGISON for treatment of Psoriasis
FOR 3 YEARS
Immigrant or refugee coming from an area endemic for Malaria, 3 years after departure, or those
who have had a Diagnosis of Malaria, 3 years after becoming asymptomatic
Have taken the drug SORIATANE for treatment of Psoriasis
FOR 1 YEAR
Sexual contact w/ a prostitute or persons in a high-risk group for AIDS
o Sexual worker
o IV drug users
Rape victims
After Hepatitis B immune globulin administration
Close contact with someone with symptomatic viral hepatitis
Tatoo, skin piercing, accidental needle sticks
Recipient of blood components thru transfusion
Recipeint of organ transplants
Treatment of syphilis/gonorrhea (deferral starts after completion of therapy)
After therapeutic rabies vaccination
After bite from an animal
Health care workers w/ percutaneous exposure to blood / body fluids
Incarceration in a jail for more than 72 consecutive hours
FOR 6 MONTHS (After cessation of the drug Avodart for prostate gland enlargement)
FOR 1 MONTH –
German measles (Rubella) vaccination
After cessation of the drug Proscar for treatment of Benign Prostatic Hyperplasia
After cessation of the drug Accutane for Acne treatment
After cessation of the drug Propecia for baldness
FOR 2 WEEKS
After vaccination with: Yellow fever, Oral polio, Measles (Rubeola), Mumps, Typhoid
TEMPORARY
Active disease under treatment: Cold, Flu, TB, Syphilis, infections
Curable disease of the heart, lung, kidney, liver, and GI tract
Treatment w/ ANTIBIOTICS
QUALIFICATIONS OF A POTENTIAL BLOOD DONOR
2. Body weight
110 lbs or 50 kg
Standards mandates a maximum of 10.5 mL/kg of donor weight for whole blood collection
inclusive of pilot tubes for testing
3. Temperature
Oral temperature should not exceed 37.5ºC or 99.5ºF
4. Pulse
50 – 100 beats / min
Lower pulse rate is acceptable for athletes
5. Blood Pressure
Systolic: < 180
Diastolic: < 100
6. Minimum Hemoglobin
Hct: > 38%
Hb: > 12.5 g/dL
Hct = 33%
Hb = 11 g/dL
Last donation must be completed 3 days before the anticipated operation
Tests to be done: ABO & Rh