Blood Groups

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ABO Sytem

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• Discovered (accidentally) in 1900 by Landsteiner
  
• Discovered 3 (A, B, C) of 4 blood groups by testing co-workers blood
• Awarded Nobel prize in 1930 for his discovery

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Antigens

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• A 43% occurrence
• B 9% occurrence
• C (now O) 45% occurrence
• AB 3% occurrence

• first development 6 week feotus; 50% at birth

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Gene to Antigen

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Front 4

Genes; FUT1 and FUT2

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• FUT1; responsible for presence on red cells
• FUT1(H) gene encodes for alpha1,2-fucosyl transferase

- Transfers L-fucose to terminal sugar membrane chain, galactose. H antigen

- Present in all O group individuals; necessary precursor for addition of A and B antigens

• FUT2; responsible for presence in tissues
• FUT2(Se) gene; presence of sugars in bodily fluids

- Endodermically derived tissues have H antigen generated by alpha1,2-l-fucosyl-transferase

Front 5

Precursor substance

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• X--Gal--GlcNac---Gal

- X; Carbon chain

- Gal; D-Galactose

- GlcNAc; N-Acetyl-D-Glucosamine

• 
  

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A, B, H Gene-Enzyme-Carbohydrate

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Front 7

Genetics; 3 separate loci

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• A, B, O Chromosome 9
  
• H active, h silent amorph; Chromosome 19
• Se active , se silent amorph; Chromosome 19

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Phenotypes and Genotypes

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• A; AA or AO
• B; BB or BO
• AB; AB
• O; OO

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Bombay Phenotype

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• H gene is absent giving hh genotype (inactive)
  

• Precursor substance is unchanged (absence of L-fucose on chain)

- therefore A/B antigens are not produced

• A/B genes inherited and transmitted normally to offspring.

• Test positive for O blood group agglutination although Anti- A, B, H antibodies present in serum.

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Antibodies

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• "Naturally acquired" - result from stimulus of non pathogenic microorganisms (ingested food)

• Form 3 month gradually increase peak at 5 years and decline with old age

• Usually form as Ig M class; also form in mixture of Ig G, Ig A

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A Subgroups

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• A1 (80%) presents A, A1 antigens
• A2 (20%) present only one form of A antigen

- A1 can be tested for using plant lecin of Dolichos Biflorus

- Subgroup A2 weak agglutination with anti-A

• A3, Ax occur infrequntly and have less antigen sites increase in H antigen activity

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B Subgroups

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• Rare (B3, Bw, Bm) higher frequency in asian and african populations.

• variable reaction with Anti- B and Anti-A,B

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Other groups

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• Acquired B
  

-Group A to AB

-Plasma reacts with anti-B

-Red cells reacts with anti-A and weakly with anti-B

- N-acetyl-D-galatosamine to galactoseamine similar to D-galactose

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Other Groups

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• CIS AB

- A/B genes on same chromosome; unequal crossing over
- Gene mutation; mutant enzyme makes both antigens

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Manual Testing

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Front 16

Automated Testing

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• Column agglutination technology
• Preloaded Micro-column cassette/card
• Glass beads/Gel matrix

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Ortho

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Front 18

Diamed

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• Gel test
• Invented by Yves Lapierre in Lyon 1984
  
• Commercial kits available from Diamed 1988

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Olympus

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• Positive test appears as a homogeneous layer of cells
• Negative test appears as a compact dense cell button surrounded by a clear zone

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Errors

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• Improper Identification of patient sample
• Incorrect result interpretation/recording
• Under/over-centrifugation
• Failure to add reagent/plasma to tube
• Failure to add red cells in appropriate concentration
• Incorrect procedure followed e.g. incubation time/temperature
• Contaminated reagents/red cells/saline or glassware
• Failure to detect rouleaux

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Controls (4)

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1. Positive Control
2. Negative Control
3. Autologous Control
4. Checkboard Record

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Rh System

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• Discovered in 1940 by Landsteiner and Wiener

• Agglutination of rabbits and guinea pigs with Macacus rhesus red cells

• Rh D factor in one system, CcEe recognised in 1945

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Rh Antigens

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• D

- Antigenic, immunogenic

- 85% population express D

- 30 different epitopes

- Weak D fewer antigen sites

- Partial D incomplete D ; missing epitopes

• C (70%) c (80%)

- Antithetic

- co-dominant genes

• E (30%) and e (98%)

- Antithetic

- co-dominant genes

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Antibodies

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• Anti-D, Anti-C. Anti-c, Anti-E and Anti-e

- Anti-D most common most immunogenic

- Followed by c, E, C (rare), e (rare)

• Class IgG
• Antibody screen (IAT)

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Fisher-Race

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• Most commonly used terminology
• Three linked genes D, C, E
• DCE now CDE

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Wiener

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• R D antigen
• r absence of D
• C indicated by 1 or (')
• c there is no 1 or (')
• E indicated by 2 or ('')
• e there is no 2 or ('')

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Phenotyping

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• Anti-sera of known specificitiy used

- Anti-D, Anti-C, Anti-E, Anti-d, Anti-c, Anti-e

• React with patients red cells

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Genetic; Tippett

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• Tippett 1986

• Two distinct genes; on chromosome 1

- RHD, RHCE

• D gene encode for D antigen
• CE gene encodes for ce. Ce, cE or CE

- co-dominant alleles

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Rh Testing

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• Tested twice for first time donors

• Use monoclonal anti-D reagent

• Control reagent (patients)

• Full Rh phenotype (donors)

• Purposed of testing;

- Prevent stimulation of anti-D antibodies
- Detect presence of Rh antibodies

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Rosenfield

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• Alpha-numeric; number assigned to each antigen;

- D: Rh1

- C: Rh2

- E: Rh3

- c: Rh4

- e: Rh5

• Presence/absence denoted by ±

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ISBT

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• International Society of Blood Transfusion
• Standarise nomenclature; 6 digits assigned
• Rh system denoted by 004

- D:004001

- C:004002

- E:004003

- c:004004

- e:004005

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Rh Antigens in red cell membrane

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• D and CE produce separate proteins
• 2 x RhAG
• Spans membrane both intra- and extracellular surfaces; loops 6 times

- NH2 and COOH termini located intracellularly

• 417 amino acids

- 36 differ between RhD and RhCE

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Other Blood Groups (6)

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1. Kell (006)
2. Duffy (008)
3. Kidd (009)
4. MNSs (002)
5. Lewis (007)
6. P (003)

** 1-4 are protein-based systems and 5-6 are carbohydrate-based systems **

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Kell System (Kel)

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• 006: ISBT numbering system
• Identified 1946 as causative antigen for Haemolytic Disease of Fetus and Newborn (HDFN)
• Antigen expressed on red cell membrane glycoprotein with membrane-bound enzyme activity (zinc endopeptidase)
• K and k genes are co-dominant; alleles on chromosome 7
• K (kell) present in <9% popuation
• k (cellano) present in > 90% population
• K antigen most immunogenic outside of ABO and Rh systems
• IgG antibodies and clinically signiicant

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Duffy System (Fy)

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• 008: ISBT numbering system
• Expressed on red cell membrane glycoprotein with chemokine receptor activity
• DARC gene - Six antigens; co-dominant alleles Fya and Fyb located on chromosome 1 q22-q23
• Fya discovered in 1950 and Fyb in 1951
• IgG antibodies; do not bind complement but clinically significant

- stimulated by transfusion and pregnancy (not common cause of HDN)

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Duffy system; population

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• Homozygous silent gene rare in Caucasians but 68% in Blacks
• Fy(a-b-) cells protected against invasion by malarial parasites

-Plasmodium knowlesi and P. vivax

• Fy antigen acts as receptor for invasion

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Kidd System (JK)

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• 009: ISBT numbring system
  
• Expresed on the red cell membrane glycoprotein which transports urea across membrane
• SLC4A1 gene located on chromosome 18 q11-q12
• Three antigens; co-dominant JKa (1951) and JK (1953)
• IgG antibodies clinically significant

- implcated in HTR an HDFN

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MNSs System

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• [43] Antigens carried on glycophorins (Sialoglycoproteins)

• MNS second blood group discovered in 1927 using IAT; red cells injected in rabbits
• co-dominant genes GYPA and GYPB on chromosome 4 (q28.2-q13.1)
• M and N antigens expressed glycophorin and S and s expressed on glycophorin B

- antigens differ amio acid sequnce on GYP

- GYP A acts as receptor for P. falciparum

• U antigen always present when S and s inherited

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MNSs antibodies

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• MN: usually IgM, cold reacting and insignif.

• M detected in saline, N is very rare

• Ss and U: all IgG and clinically significant

• S and s produced by allelic antigens at locus closely inked to MN locus

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Lewis (Le)

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• Antigens are not intrinsic to RBC; produced by secretory tissue and absorbed from the plasma

• Major antigens Lea (1946) and Leb (1948)

• Encoded for by Le or (FUT3) gene on chromosome 19

- interacts with Se and H genes

- A, B, H AND Le secretions

• Antibodies naturally occurring, usually IgM and not clinically significant

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P Blood Groups

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• Discovered in 1927 when Landsteiner and Levine immunised rabbits with RBCs

- Initially named P, renamed to P1

- RBCs lacking P1 and P2

• P antibodies are usually IgM and react optimally at 4 degrees C

• P1 antigen diminishes in storage and strength varies among samples.

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Summary Other Blood Groups

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•Antigens– RBCtransmembrane proteins

•Antibodies–Variableclinical significance

•Terminology–Traditionaland ISBT

•BGS–Kell,Duffy, Kidd, Lewis, MNS, P