ABO Blood Group System

• The presentation is on blood banks and was presented by Haya Mansour for the SLLE Exam 2023.

Topics Covered

• Major Blood Groups Systems
• Minor Blood Group System
• ABO Discrepancy
• Antibody Screening
• Antibody identifications
• Techniques used in blood bank
• Hemolytic disease of the newborn HDN
• Blood donation
• Blood components

Major Blood Group Systems

• ABO blood group system
• Rh Blood Group System

ABO Blood Group System

• It has one position, or locus on each chromosome 9 consisting of an A, B, or O gene.
• Each person has one blood group of: A, AB, B or O.
• These entities are carbohydrates attached to proteins or lipids (glycoproteins, glycolipids).
• They are called antigens when they interact with another body.
• They are called proteins when they are inside the body as part of RBC structure.
• ABO Abs are mostly IgM class

Important Systems

• These contain antigens can provoke the most severe transfusion reactions:
• ABO system first described by Karl Landsteiner in 1900.
• Rhesus system.
• Other blood group systems, such as Kell, Kidds, Duffy etc.

ABO System Antigens

• They are normally present as solid on RBCs.
• Also in the plasma, saliva, semen, and other secretions.
• ABO antigens are not found in CSF.
• ABO antigens are also expressed on various body tissues.
• They exist on T, B cells and platelets when they adsorbed from the plasma.

Antigen Importance

• They are the most immunogenic.
• Cause the most common cause of death from a blood transfusion.
• ABO blood types vary among different populations.
• ABO antigens are important in organ transplantation.

Inheritance of ABO Antigens

• One A,B,O gene is inherited from each parent
• A genes are dominant
• B genes are dominant
• A & B genes are Co- Dominant
• O gene is recessive

Phenotypes and Genotypes

• A phenotype has an AA or AO genotype
• B phenotype has an BB or BO genotype
• AB phenotype has an AB genotype
• O phenotype has an OO genotype
• The phenotype is the result of the reaction between the red cells and antisera.
• The genotype is the genetic makeup and can be predicted using the phenotype.
• A group A individual has a AA/AO genotype.
• If the father is A and the mother is O and the baby is O then the father must be heterozygous.

ABO Types

• O is the Universal donor
• AB is the Universal recipient

Frequencies of types

• In whites:
  • O is observed 45% of the time
  • A is observed 40% of the time
  • B is observed 11% of the time
  • AB observed 4% of the time
• In blacks:
  • O is observed 49% of the time
  • A is observed 27% of the time
  • B is observed 19% of the time
  • AB observed 4% of the time

ABO Blood Rules

• A person does not have antibody to his own antigens.
• Each person has antibody to the antigen lacks only in the ABO system

Subgroups A and B

• Subgroups of A and B antigens:
  • They are inheritance conditions.
  • This subgroup has fewer H chains converted to A or B antigen.

Subgroups of A antigen

• A1, A2, A3, Ax, A.int, A.m, A.el, A1B, A2B.
• 80% of group A people are A1.
• 20% of group A people are A2.
• A1 antigen has five times sites more than on A, and few Η antigen.
• A2 has more free H antigen and antibodies anti-A1 that is usually clinically insignificant.

Subgroups of B antigen

• B3, Bx, B.m, B.cl

Formation of Red Cell Antigens

• ABH genes code for produce specific glycosyltransferases that add sugars to a basic precursor substance.
• A, B, and H antigens are formed from a basic precursor material (called glycan) to which sugars are attached in response to specific enzyme transferases.
• The H antigen is the precursor of A and B antigens.
• The H gene results in the formation of the Η antigen, Η and Se genes influence A and B antigen expression.
• The H gene must be inherited to form the ABO antigens on the RBCs, and the Se gene must be inherited to form the ABO antigens in secretions.
• When L-fucose sugar attaches to an oligosaccharide chain on the terminal galactose of type 2 chains, a H substance is formed.
• When N-acetyl-D-galactosamine sugar transfer to the H substance the blood group A is formed.
• When a D-galactose sugar transfers to the H substance, the blood group B. is formed
• Genotype hh (Bombay), the production of a-2-L-fucosyltransferase, causes there to be no L-fucose, no A, B, O Ags produced.

Donor Nucleotides & Immundominant Sugars

• Gene H with the Glcosyltransferase L- fucosyl trnsferas, that adds L-fucose to create H
• Gene A with the Glcosyltransferase N acetylglucosaminyl transferase, that adds N-acetyl D to create A
• Gene A with the Glcosyltransferase D galactosyl transferase, that adds D-galactose to create B
• H: O > A2 > B > A2B > A1 > A1B

ABO-Antibodies Characteristics

• They Form naturally, they arise without immune stimulation.
• IgM for A and B blood group.
• IgG for blood group O.
• They can fix the complement.
• Antibodies exist in the plasma.
• Absent at birth or they are very weak. The're not detectable in the blood of newborn infants due to their underdeveloped immune system Antibodies start to appear at age 4 months; stable by ages 5-6 years and reach adult levels at age 10,

Antibody Definitions

• Naturally: formed without prior exposure for foreign antigen.
• Autoantibody: antibody formed against patient own antigens.
• Alloantibody: formed to foreign antigens.
• Anti-A and/or anti-B present at birth are of maternal IgG origin.
• May disappear with elderly age.
• Older age or Bone marrow transplantation.

Blood Group Antibodies

• Can cause:
  • Haemolytic transfusion reaction (HTR) because of ABO-mismatched blood transfusion
  • New born Haemolytic transfusion reaction in the embryo A or B the mother O blood group.
  • Acute graft rejection because of ABO-incompatible solid organ transplantation
  • Haemolysis of donor red cells because of ABO-incompatible bone marrow transplantation.

Lectins and Subgroup A

• Lectins are extracts of seeds of plants that react specifically with certain antigens on the RBC.
• They act like antibodies.
• Lectins in blood banking:
  • Ulex europaeus, Lectin H agglutinates RBC cells that have Η substance.
  • Dolichos biflouros, lectin Al, agglutinates RBC cells with A1.
• Lectin-H - strongest reaction with O cells, high concentration of Η antigen and weakest with A,cells, have a low concentration of H

Lectin Reactions

• Lectin-A reacts strongest with Al cells and A, B cells which has a low concentration of H antigen.
• Negative with the other RBC cells with high concentration of H.
• Al: has 4+ Anti-A, 4+ Anti-A1, 0 Anti-H Lectin, Anti-B antibodies in serum
• A2: has 4+ Anti-A, 0 Anti-A1, 3+ Anti-H Lectin, Anti-B and Anti-A1 antibodies serum

Bombay Phenotype

• The designation "Bombay" phenotype was assigned to people who lack A, B, and H antigen
• Η antigen is not expressed on RBCs nor found in saliva
• In the Genotype: h/h se/se there's no production of a-2-L-fucosyl-transferase, has no fucose and lack of A, B, O Ags
• Referred to as having the "Oh“ and Represent <0.01% of the population.
• A blood test is typed as O blood group.
• serum contains anti-H.
• On cross-matching, all units are incompatible.
• The serum contains anti-A, anti-B, and anti-H antibodies
• All the antibodies are IgM.
• ALL are naturally occurring.

Para-Bombay Phenotype

• null alleles
• Η antigen is weakly expressed on RBCs.
• There may be or not be saliva.
• Genotype: (H), Se/Se or Se/se
• Normal amount of H antigen is detectable in saliva
• Requires H-negative blood (Bombay donors)
• Serum Contains anti-H,

Bombay and Para-Bombay

• If ABO bloods tests type as group O
• Antibody - Plasma contains strong anti-H with anti-A and anti-B
• Cross-matching - Serum incompatible with O cells
• anti-H lectin - Red cells yield negative reaction

ABO Typing

• Definition - a test to targeted to:
  • Detect Antigens on cells (RBC).
  • Detect antibodies in serum or plasma

Types of ABO tests

• Forward typing (cells grouping) - detect ABO antigens on the RBC surface for donors and recipients
• Reverse typing (serum typing) - detect ABO antibodies in serum or plasma of control RBC and recipients

Detection of Antigen-Antibody

• The detection of antigen-antibody reactions in the blood banking has two possible forms:
  • Hemolysis-lysis or destruction of RBC with the formula Ag+ Ab + C'=lysis of RBC
  • Agglutination - clumping of RBC in the presence of Ab with the formula Ag + Ab = clumping of RBC

Agglutination Technology

• Gel or column agglutination technology: - with positive results, Cells remain at the top of the column
• with negative results, Cells settle at the bottom

ABO Discrepancies

• Occur with unexpected reactions in the forward and reverse grouping. Note that the RBC and serum grouping reactions that are 3+ or 4+ are very and weaker reactions represent discrepancy.
• The problem is:
  • Missing reaction
  • Weaker reaction
  • Extra reaction

Resolving

• To resolve: -. Eliminate technical errors.
  • Enhance the reaction in reverse grouping.
  • Incubate the patient's serum with reagent cells at room temperature for 15 minutes.

Possible Discrepancies

• Group I discrepancies - Missing antibodies - found most often in in new born infants and elderly patients plus Patients with lymphoma, using immunosuppressive drugs, with ABO subgroups and with immunodeficiency disease/BM transplant
• Group II discrepancies - Missing antigens - caused by some subgroups of A or subgroups of B or present in patients with leukemia and Hodgkin's disease.
• Group III discrepancies - these discrepancies between forward and reverse grouping due to protein abnormalities caused by elevated levels of globulin from certain diseases including multiple myeloma, Hodgkin's lymphoma and Rouleaux formation.
• Group IV discrepancies - Between forward and reverse groping due to miscellaneous problems including bacterial or viral infections for Bone marrow and Stem cell recipients

Fixing Discrepancies

• Group II discrepancies wash the patient's cells with saline.
• Group III discrepancies adding a drop or two of saline to the tube in case of rouleaux formation and the red cell clumping will remain the same for true cases.

Grading Reactions

Grading from Negative reaction, up to 4+ Reaction, or Hemolysis.

Anti-A, Anti-B, and anti-AB

• The anti-A, anti-B, and anti-AB in the reverse grouping test can be used to determine a blood group

Rh blood group system

• It's a Complex blood group system with around 50 different antigenic specificities
• Is Co-dominant and found on the Chromosome 1.
• Rh antigen is a Rh protein
• Expressed only on RBCs and well developed at birth.
• It is made of Two closely linked genes, Rh D and Rh CE.
• Five antigens form the system, D, C, с, Е, е
• Rh antigens are not soluble.
• Only Rh antibodies are produced after exposure to foreign red blood cells in HDN
• D antigen is highly immunogenic and the primary cause of hemolytic disease of the fetus and newborn.

Rh and Antibodies

• The Rh antigens are highly immunogenic but the D antigen is most potent.
• IgG is needed to become sensitized to red blood cells (HDN).
• Rh antibodies are produced only after exposure to foreign red blood cells.

RHD locus

• One antigen called D, is Dominant, expresses all the D antigen epitopes.
• RhD positive calls the D antigen present while if not there, RhD negative
• D is a strong antigen and extremely immunogenic
• It Is the most important in clinical practice

RHCE locus Characteristics

• Contains two antigens C/c and E/e, which are co-dominant an express themselves at 4 allelic forms Ce, ce, cE or CE
• One protein (RHCE) carries both C/c or and E/e antigens.
• Combination determined by which alleles of RHCE are present (CE, Ce, cE, or ce).

Frequency

• In whites, in order of most to least frequent: e, D, C, E,c
• In blacks, in order of most to least frequent: e, D, c, E, C

Terminology

• Is based on the theory that antigens produced by three closely linked sets of alleles and each gene is responsible to produced (or antigen) on the RBC surface, and consists of D, d, C, c, and E, e where “d" is absence of D Ag. -Homozygous has positive Rh -Heterozygous has positive Rh -Homozygous has negative Rh
• Anti D antiserum is used to determent D positive or negative

Weiner Theory

• Wiener theory rh-hr terminology is the concept that there was one gene complex with varying alleles and that various Rh antigens are produced • R means the presence of the antigen "D" whiler indicates the lack of it • Presecne of C or E is indiacet by 1'' or 2'' presence, and the lack, by by empty terms • presence of both is shown by Rz or Ry
  • if r precedes h and starts with C or E, antigens must written in "Capital letter terms"
  • if h precedes r, and ends with c or e, antigens murt be written in "small letter terms "

Wiener to Fisher-Race

• Fisher-Race, - Weiners systems' main differences lies in Weiners major idea of 1 genes producing multiple antigens, versus Fishers Race's 3 allele sets each producing a singular trait
• Translate Wieners to Fischer race as such:
• R refers to D.
• Rº, R1, R2, and Rz. → presence of D Rh positive).
• r,r',r", and r →Absence of D (Rh negative).
• Rº → c & e.
• R¹ → C
• R2 → E
• Rz → C & E.

Rh-Hr Terminology of Wiener

• Has shorthands for antigens and genes following this system:
• Rho shorthanded to Ro indicates Dce
• Rh₁ shorthanded to R₁indicates DCe
• Rh² shorthanded to R₂ indicates D.cE
• Rh shorthanded to Rz indicates DCE
• rh shorthanded to r is ce
• rh' shorthanded to r'is Ce
• rh" shorthanded to r" is cE
• rhy shorthanted to ryIs CE

Rosenfield Terminology

• Number to each antigen of Rh system, no genetic basis but simply demonstrates the presence or absence of Ags on the red cells where:
  • Rh 1 = D
  • Rh 2 = C
  • Rh 3 = E
  • Rh 4 = c
  • Rh 5 = e
• In writing the phenotype, the prefix "Rh" is followed by colon, then number (preceded by negative sign), e.g. D+, C+, E-, c+, e+ is written as Rh:1,2,-3,4,5.

Common Types by Three Nomenclatures Table

Common genotypes:

• R1r/ DCe/dce is Rh:1, 2, -3, 4, 5
• R1R1/ DCe/DCe is Rh:1, 2,3, -4,5
• rr/ dce/dce is Rh:1, -2, -3, 4, 5
• R1R2/ DCe/DcE is Rh:1, 2, 3, 4, 5
• R2r /DcEdce is Rh:1, -2, 3,4, -5 Rarer genotypes:
• R2R2/ DcE/DcE is Rh: 1, -2, 3, 4, -5
• r'r/ dCe/dce is Rh:-1, 2, -3, 4, 5
• r'r'/ dCe/dCe is Rh: -1, 2, -3, -4, 5
• r"r/dcEdce is Rh:-1, -2, 3, 4, 5
• r"r'/ dce/dCe is Rh:-1, -2, 3, 4,-5
• Ror RORO Rar (RORO) = Rh:1, -2, -3, 4, 5
• I'r"r" (ryr) is Rh:-1, 2, 3, 4, 5

Uncommon Phenotypes and Weak D

• Inherited
• Genetic weak D: has reduced antigen number on the cells
• Not expressed as D negative, no anti-D
• Received D positive and negative blood
• Partial D, missing components
• Can only receive Rh neg blood and be demonstrated by the Coombs technique as “AGT"

Minor Blood Group Systems

• Lewis
• I
• P
• MNSS
• Kell
• Kidd
• Duffy
• Lutherean.

Terms

• Clinical significance: antibodies associated with decreased RBC survival: Transfusion reactions and HDN.
• Not clinically significant will not the red cells
• Cold reacting antibodies: agglutination best observed at or at or below room temp. Warm reacting antibodies: agglutination best observed at 37°C

Cold-Reacting Antibodies

• Lewis antigens
  • Soluble antigens produced by tissues, in body fluids (plasma).
  • Adsorbed on the RBC
  • Lewis substance adheres to RBC
  • Become an antigen RBC Le substance with plasma Le genes.
• Lewis antibodies
  • Anti-Lea and Anti leb.
  • are Is IgM.
  • No clinical significance.
  • Pregnant women (a-b-).

Produce Cold-Reacting Antibodies and I Antigens

• These antigens may be i or I, form on the precursor chain of RBCs
• Newborns have i antigens, adults have I antigen with linear antigen in newborns and branch as they age
• The antibodies are Is IgM antibody found in normal adults, NO Clinically significant and associated as Cold Agglutinin Disease and Mycoplasma pneumoniae
• Anti-i antibodies: are Rare and associated with infectious mononucleosis

Produce Cold-Reacting Antibodies and the P Antigen

• Most common phenotypes are P1 and P2
  • where P1 - consists of these two,while, P2 - consists of only P antigens.
• 75% of adults have P1 group and are able to produce anti-P1
  • The combination Includes Anti-P, anti-P1, and anti-Pk
  • Found in serum of the people who do not have any of those anti-P, anti-P1, and anti-Pk and anti-P antibodies

MNSs Antibody Characteristics

• Has 4: M, N, S, s, U
• M & N located on Glycophorin A, and is common
• M & N give a stronger reaction when homozygous (M+N-) or (M- N+):
• Weaker reaction occur in the heterozygous state (M+N+):
• Destroyed by enzyme and located on Gylycophorin B.
• Antibodies may be IgM or IgG

Produce Warm-Reacting Antibodies

• Kell system (Kell and Cellano):
• Well developed at birth.
• K antigen is very immunogenic.
• Kell antibodies:
  • Is IgG that is causes HTR or HDN.
  • Clinically significant.
  • Produced as a result of immune stimulation.
  • Some rare antibody in IgM

Kidd system:

• Kidd Antigen,
  • Have 2 antigens, always paired, Well developed at birth.
  • Anti-Jka and Anti-Jkb are clinically significant and cause DHTR or HDN
• Anti- JK3: present in some JK(a-b-).

Duffy system

• 2 Antigens (Fya and Fyb) exist and Are well developed at birth
• IgG. antibodies Clinically and Stimulated by transfusion or even pregnancy, known as HTR, Delayed Hemolytic Transfusion Reaction.
• Westen African phenotypes (Fya/b-) resist P.vivex by preventing malaria.

Lutheran system:

2 antigens (A and B) with 92% population having the Lu b antigen and Lu(a-b+) is "common'" but still RARE

• The antibodies are anti-Lu a, anti-Lu b, - some mild in IgM but Clinically significant IgG that transfusion reactions.

Red Cell Dosage Phenotypes

High Vs Low Incident Antigens

• High incidencent, Ags that occur in at least 98% of the population, cause big challenge to crossmatch
• Low incident Ags not present, cause negative antibody and a positive cross match

Identification

• Use Dithiothreitol, and or ZZAP

Human Leukocyte Antigen (HLA)

• A protein on molecules on cell membrane and part of the the immune system, located on chromosome 6 and Divided at 3 classes:
• Transmembrane glycoproteins present on all nucleated cells
• Class II expressed in immune cells only
• Immune response incompatible transfusion, organ tissue
• Can cause HLA must in tested, used for organ transplantation

Plate Let Antigens

• Membranes have protein antigens who can create Platelet antibodies
• Can result in Neonatal alloimune thrombocytopenia, and Potstransfucion purpurae

Cross Matching

• Cross matches test Compatibility of of a donor with that recipient
• Major cross match
  • Detect the patients abs against the donor cells
  • minor test the patients abs against the donors
• No agglutination: compatibility- agglutination: incompatible

Coombs Test

• Detect Antiforgin bodies, with to reagents, polyspefici( and monospecific
• There are two types
• "DAT" Direct Antiglbin test
• • used on In- Vivo
• "ITA" Indirect, used invitito.

DAT procedure

• Add AHG after the solution and wash
• Red cells coated with antibodies
• Will detect HDNS etc

ITA procedure

• Patient serum and drops of RBC
• After, add ahg etc
• Detects Crossmatch

Antibody Screening

• The aim to identify antibodies other those of Anti A or Anti B
• The antibodieas,
• It is very rare, screened for all important

Antibody Identification

Conduction for, and result in identifying the exact types of antibodies who create a positive reaction.

Techniques Used in the identification of antibodies

• ELIT technique The free types of any antibodies is confirmed through sensitized test

To reduce allosobties from any sample, using a serum

Prewoarmed Test

• Heat the cells to react under heat
• So use to find GG anti bodies which only respond to it.

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Transfusions Reaciton

• Types of the:
  • Heatmolic, due to transfusion
  • Ananphylatic, due to allergic transfusion
  • Heat related reastions

Donatiion:

For donation all those are needed 18 y, weight 50 kg. To donate with temp, and blood presure

Donar, deferals:

Must not do drugs: Must not do drugs Have symptoms

Blood Componets

Componets with, rbr, plet, and p

Coagualtes

Cpds and sags