Drmsaiem Rh Blood Group System DR. MOHAMMED H SAIEMALDAHR BB Faculty of Applied Medical Sciences MED...

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Rh Rh Blood Group SystemBlood Group System

DR. MOHAMMED H DR. MOHAMMED H SAIEMALDAHRSAIEMALDAHR

BBBBFaculty of Applied Medical Faculty of Applied Medical

SciencesSciencesMED TECH DEPMED TECH DEP

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Rh Blood Group SystemRh Blood Group System

The term The term RhRh refers not only to a specific red cell refers not only to a specific red cell antigen antigen

but also to complex blood group system that is currently but also to complex blood group system that is currently comprised on more than 50 different antigenic comprised on more than 50 different antigenic

specificities.specificities.

History of the Rh SystemHistory of the Rh System

Levine and Stetson described a hemolytic transfusion Levine and Stetson described a hemolytic transfusion reaction in an obstetric patient following delivery of reaction in an obstetric patient following delivery of stillborn infant. The women required transfusion. Her stillborn infant. The women required transfusion. Her husband, who had the same ABO type, was selected as husband, who had the same ABO type, was selected as her donor, after transfusion the recipient, her donor, after transfusion the recipient, demonstrated the classic symptoms of acute demonstrated the classic symptoms of acute hemo]ytic transfusion reaction. hemo]ytic transfusion reaction.

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Subsequently an antibody was isolated from the Subsequently an antibody was isolated from the mother’s mother’s

serum that react both at 37°C and 20 °C with the father’s serum that react both at 37°C and 20 °C with the father’s

red cells. It was postulated that the fetus and the father red cells. It was postulated that the fetus and the father

possessed a common factor that the mother lackedpossessed a common factor that the mother lacked..

While the mother carry the fetus, the mother was While the mother carry the fetus, the mother was

exposed to this factor and subsequently built up an exposed to this factor and subsequently built up an

antibody that reacted against the transfused red cells from antibody that reacted against the transfused red cells from

the father and resulted in hemolytic transfusion reaction.the father and resulted in hemolytic transfusion reaction.

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Landsteiner and Wiener reported on an antibody Landsteiner and Wiener reported on an antibody by guinea pigs and rabbits when they were by guinea pigs and rabbits when they were transfused with rhesus monkey red cells.transfused with rhesus monkey red cells.

This antibody which agglutinated 85% of human This antibody which agglutinated 85% of human red cells was named Rh.red cells was named Rh.

The name Rh was retained for the human The name Rh was retained for the human produced antibody.produced antibody.

Anti-rhesus formed by the animals was renamed Anti-rhesus formed by the animals was renamed anti-LW (Landsteiner and Wiener).anti-LW (Landsteiner and Wiener).

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Nomenclature of the Rh SystemNomenclature of the Rh System

The terminology used to describe the Rh system is The terminology used to describe the Rh system is derivedderived

from from 4 4 sets of investigators.sets of investigators.

TwoTwo of the terminologies are based on the of the terminologies are based on the postulated postulated genetic mechanisms of the Rh systemgenetic mechanisms of the Rh system..

33rdrd terminology describes only the terminology describes only the presence or presence or absence of a given antigen.absence of a given antigen.

44thth is result of the effort of the is result of the effort of the International International Society of Blood Transfusion (ISBT)Society of Blood Transfusion (ISBT) Working Working Party on Terminology for Red cell Surface Party on Terminology for Red cell Surface antigens.antigens.

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Fisher-Race (DCE terminology)Fisher-Race (DCE terminology)

In the early 1940s, Fisher and Race were In the early 1940s, Fisher and Race were investigating investigating

the antigens found on human red cell including the the antigens found on human red cell including the newly newly

defined Rh antigen.defined Rh antigen.

Thy postulated that the Thy postulated that the antigensantigens from the system from the system werewere

produced by produced by 3 closely linked sets of alleles3 closely linked sets of alleles. Each . Each genegene was responsible for was responsible for producing a productproducing a product ((AgAg) on the red cell surface.) on the red cell surface.

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Each Each antigen (Ag) and corresponding geneantigen (Ag) and corresponding gene were were given the same given the same letter designationletter designation. Fisher and Race . Fisher and Race named the antigens of the system named the antigens of the system D, d, C, c, E, and D, d, C, c, E, and e.e.

To date To date NONO ( (d ) d ) antigen has been found, and it is antigen has been found, and it is considered an considered an amorph (silent allele)amorph (silent allele) or the or the absence of absence of DD antigen. antigen.

The phenotype (blood type observed during The phenotype (blood type observed during testing) of a given red cells is testing) of a given red cells is define by the define by the presence or absence of D, C, c, E, and e.presence or absence of D, C, c, E, and e.

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According to According to the Fisher-Race proposal, each the Fisher-Race proposal, each person inherits a set of Rh genes from each person inherits a set of Rh genes from each parent (one D, or d, one C or c, and one E or parent (one D, or d, one C or c, and one E or e).e).

Rh genes are Rh genes are co-dominantco-dominant, each inherited , each inherited gene express its corresponding antigen on gene express its corresponding antigen on the red cell.the red cell.

The combination of maternal and paternalThe combination of maternal and paternal haplotypeshaplotypes ( (Possessing half the normal # of chromosome found on Possessing half the normal # of chromosome found on

the cellthe cell) ) determines one’s determines one’s genotypegenotype (the Rh gene (the Rh gene inherited from each parent) and dictates one’s inherited from each parent) and dictates one’s phenotypephenotype (the antigens expressed on the red cell (the antigens expressed on the red cell that can be detected serologically).that can be detected serologically).

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An individual’s Rh phenotype is reported as An individual’s Rh phenotype is reported as DCEDCE because Fisher-Race postulated that the because Fisher-Race postulated that the C/c C/c locuslocus is lies between is lies between D/d D/d and and E/eE/e loci. loci.

It is essential to remember that It is essential to remember that dd doesn’t doesn’t represent an antigen but simply the represent an antigen but simply the absence of absence of the the D antigenD antigen. .

C, c, E, e C, c, E, e represent actual represent actual AgsAgs recognized by recognized by specific antibodies.specific antibodies.

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2- 2- Wiener (The Wiener (The Rh-HrRh-Hr terminology) terminology)

Wiener believed that, the Wiener believed that, the gene responsible for gene responsible for defining Rh actually produced an agglutinogendefining Rh actually produced an agglutinogen ( (a a substance that stimulates the production of an substance that stimulates the production of an

agglutininagglutinin, thereby acting as an , thereby acting as an antigenantigen)) that that contained a series of blood factors.contained a series of blood factors.

The The agglutinogenagglutinogen may be considered the may be considered the phenotypic expression of the haplotype.phenotypic expression of the haplotype.

Each Each factor is an antigen recognized by an factor is an antigen recognized by an antibodyantibody..

Antibody can recognize a single or multiple Antibody can recognize a single or multiple factors (factors (AgAg))

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Agglutinogen Agglutinogen Red cell surfaceRed cell surface

Rh geneFactor Rh0Factor hr′Factor hr″

RH0hr ′hr ″


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Wiener (Wiener (The Rh-Hr terminologyThe Rh-Hr terminology))Fisher-Race nomenclature may be converted to Fisher-Race nomenclature may be converted to Wiener nomenclature and vice versa.Wiener nomenclature and vice versa.

It is important to remember that an agglutinogen It is important to remember that an agglutinogen in the Wiener nomenclature actually represent the in the Wiener nomenclature actually represent the presence of a single presence of a single haplotypehaplotype composed of three composed of three different antigensdifferent antigens..

R= DR= D antigen antigen rr = absence of the = absence of the DD antigen ( antigen (dd).).C=C= indicated by indicated by 1 1 or a single or a single ( ′ )( ′ ) c=c= when there is when there is no 1 or no 1 or (′)(′)EE=is indicated by =is indicated by 22 or ( or ( ″ ″ ) ) e=e= there is no 2 or ( there is no 2 or ( ″″ ) )

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ExampleExample

RR11 = = DDCCe e RR00 = =DDce ce rr′′ = =ddCCe e

E=is indicated by 2 or ( ″ ) e= there is no 2 or ( ″)E=is indicated by 2 or ( ″ ) e= there is no 2 or ( ″)

ExampleExample

R2=DcE r″ =dcER2=DcE r″ =dcE

When both C and E are present the letter When both C and E are present the letter Z or YZ or Y is used. is used.RRZZ=DCE r=DCE rYY=dCE =dCE

R1r =DCe/dce r′r =dCe/dceR1r =DCe/dce r′r =dCe/dce

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Rh Blood Group System (Wiener Rh)Rh Blood Group System (Wiener Rh)

When referring to the Rh Ag (factor) in wiener When referring to the Rh Ag (factor) in wiener nomenclature the nomenclature the single primesingle prime ( ( '' ) refers to either ) refers to either C or c.C or c.

The The double primedouble prime ( ( ″ ″ ) refers to either ) refers to either E or e.E or e.

If the If the rr precedes the hprecedes the h ( ( rh′ or rh″rh′ or rh″ ) we are referring to ) we are referring to C or C or E Ags respectively.E Ags respectively.

When the When the h precedes the rh precedes the r we are referring to either we are referring to either c c

( hr′ ) or e ( hr″ ) Ags.( hr′ ) or e ( hr″ ) Ags.Rho = D Rho = D

There is no designation for the absence of D Ag.There is no designation for the absence of D Ag.

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Rosenfield (Alpha numeric terminology)Rosenfield (Alpha numeric terminology)

In 1960s Rosenfield proposed a system that In 1960s Rosenfield proposed a system that assigns a assigns a number to each antigen of Rh systemnumber to each antigen of Rh system in in order of its discovery or order of its discovery or recognized relationship recognized relationship to the Rhto the Rh system. system.

This system has This system has no genetic basisno genetic basis but simply but simply demonstrates the demonstrates the presence or absence of the Agspresence or absence of the Ags on the red cells.on the red cells.

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RosenfieldRosenfield

A minus ( - ) signA minus ( - ) sign preceding a number designated preceding a number designated the the absence of the Ag.absence of the Ag.

If Ag has not been typed forIf Ag has not been typed for, its number will not , its number will not appear in the sequence .appear in the sequence .

D is assigned =Rh1D is assigned =Rh1 C=Rh2C=Rh2E=Rh3 c =Rh4 e = Rh5E=Rh3 c =Rh4 e = Rh5

For cell that typed For cell that typed D+ C+ E+ c- e-D+ C+ E+ c- e-, the , the Rosenfield designation is Rosenfield designation is Rh:1,2,3,-4,-5.Rh:1,2,3,-4,-5.

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RosenfieldRosenfield

If the sample was not tested for If the sample was not tested for ee, the , the designation would be designation would be Rh:1,2,3,-4Rh:1,2,3,-4..

All Rh system antigens have been assigned a All Rh system antigens have been assigned a number. number.

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International Society of Blood TransfusionInternational Society of Blood Transfusion

Numeric terminologyNumeric terminology

They adapted a They adapted a six digit numbersix digit number for each blood for each blood group.group.

The first The first three numberthree number represent the represent the systemsystem and and the remaining the remaining three represent the antigenic three represent the antigenic specificityspecificity..

The number The number 004004 was assigned to the was assigned to the Rh bloodRh blood group system, then each group system, then each antigen assignedantigen assigned to the to the Rh systemRh system was given a unique number to was given a unique number to complete the six digit computer numbercomplete the six digit computer number..

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Biochemistry of Rh AntigensBiochemistry of Rh Antigens..

Understanding the results of gene actionUnderstanding the results of gene action

The final result of gene action in red cell group is the production of the biochemical structure.

Rh system is a non-glycosylated protein this mean that,

No carbohydrates attached to the proteinRh Ags are transmembrane polypeptide (integral part of the red cell membrane)D associated protein is linked to the membrane skeleton.

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Biochemistry of Rh Antigens.Biochemistry of Rh Antigens.

The number of D Ag sites, measured on a variety of Rh The number of D Ag sites, measured on a variety of Rh phenotypephenotype

Rh phenotypeRh phenotype# of D antigen sites# of D antigen sites

RR11rr990,0-146,00990,0-146,00

RR00rr120,00-200,00120,00-200,00

RR22rr140,00-166,00140,00-166,00

RR11RR11145,00-193,00145,00-193,00

RR11RR22230,00-310,00230,00-310,00

R2R2R2R2158,00-333,00158,00-333,00

D--D--110,000-202,000110,000-202,000

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Biochemistry of Rh AntigensBiochemistry of Rh Antigens..

The greatest number of The greatest number of D Ag site are on cells of D Ag site are on cells of the rare Rh phenotype D --.the rare Rh phenotype D --.

D--D-- cell carry only cell carry only D Ag and completely lack C c D Ag and completely lack C c and E e.and E e.

R2R2R2R2 cells possess the cells possess the largest number of D Ag largest number of D Ag sitessites..

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Detection of Rh Detection of Rh Antibodies and AntigensAntibodies and Antigens

Are Are IgGIgG, react optimally at , react optimally at 37 37 ooCC or following the or following the addition of addition of antiglobulin reagentantiglobulin reagent

Produced after exposure of the individual’s Produced after exposure of the individual’s immuneimmune system to foreign red cells, either system to foreign red cells, either through through transfusion or pregnancy.transfusion or pregnancy.

Rh AbsRh Abs often persists in the circulation for years often persists in the circulation for years

Rh Abs are considered Rh Abs are considered clinically significantclinically significant, , therefore, therefore, Ag negative blood must be provided to Ag negative blood must be provided to any patient with a history of Rh Ab sensitization.any patient with a history of Rh Ab sensitization.

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Detection of Rh antibodies and AntigensDetection of Rh antibodies and Antigens

Rh Abs do not bind complement. For Rh Abs do not bind complement. For complement to be fixed, complement to be fixed, two IgGtwo IgG molecules must molecules must attach in close a proximity on the red cell attach in close a proximity on the red cell surface.surface.

Red cell destruction due to Rh Abs is primarily Red cell destruction due to Rh Abs is primarily extra-vascular. This type of hemolysis classically extra-vascular. This type of hemolysis classically characterizes characterizes a delayed hemolytic transfusion a delayed hemolytic transfusion reactionreaction..

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Rh AntigensRh Antigens

Rh antigens are Rh antigens are highly immunogenichighly immunogenic, the , the DD antigen isantigen is most potentmost potent DD c c E E C C ee

Highly Rarely Highly Rarely ImmunogenicImmunogenic

Exposure to less than Exposure to less than 1 ml of Rh positive1 ml of Rh positive red red cells can stimulate cells can stimulate Ab productionAb production in an Rh in an Rh negative personnegative person..

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Determination of D StatusDetermination of D Status

Is essential when test Is essential when test donor blood sampledonor blood sample..

Blood considered Blood considered Rh positive if either the Rh positive if either the DD or or DDuu

test is positivetest is positive

If any donor blood sample that types If any donor blood sample that types RhRhoo(D) (D)

negativenegative by either slide or rapid method must be by either slide or rapid method must be

tested further by tested further by indirect anti-globulin test (IDAT).indirect anti-globulin test (IDAT).

If both test results are If both test results are negativenegative, the donor sample , the donor sample

is considered is considered Rh negativeRh negative..

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Determination of DDetermination of Duu Status Status

There are There are instance when an accurate Rh type can instance when an accurate Rh type can not benot be determined through routine testing determined through routine testing

1- If the 1- If the new-born’s cell are coated with maternal new-born’s cell are coated with maternal IgGIgG anti-D in uteroanti-D in utero, very few D Ag sites will be , very few D Ag sites will be available to react with reagent anti-D.available to react with reagent anti-D.

Elution of the sensitizingElution of the sensitizing Ab (removing the Ab) Ab (removing the Ab) and identifying it as anti-D will verify that the and identifying it as anti-D will verify that the infant’s red cellinfant’s red cellare D positiveare D positive

2-2- Warm autoimmune hemolytic anemiaWarm autoimmune hemolytic anemia, Abs are , Abs are directed directed against the patient’s own red cellagainst the patient’s own red cell and and react as though they are react as though they are Rh specificRh specific..

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Variation of the Rho (D) AntigenVariation of the Rho (D) Antigen

Weak D overviewWeak D overviewWhen When Rh-positiveRh-positive red cell samples are typed for red cell samples are typed for the the DD Ag. It is expected that they will react Ag. It is expected that they will react strongly with strongly with anti-Danti-D reagent.reagent.However, with certain red cells the testing must However, with certain red cells the testing must be carried through the be carried through the AHG phaseAHG phase to to demonstrate the demonstrate the presence of the D Ag.presence of the D Ag.

Red cells carrying the weaker D Ag have been Red cells carrying the weaker D Ag have been referred to as having the Dreferred to as having the Duu type type..

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Three different mechanisms have been described Three different mechanisms have been described that can explain the that can explain the weakened weakened expression of the expression of the D AgD Ag..

Genetic weak DGenetic weak DInheritance of Inheritance of D genesD genes that code for a that code for a weakened weakened expression of the D Ag.expression of the D Ag.

The The D AgD Ag expressed appear to be expressed appear to be complete but complete but few in number.few in number. Inheritance of these gene can be Inheritance of these gene can be tracked from one generation to the next and seen tracked from one generation to the next and seen most frequently in most frequently in blackblack..

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C TransC Trans (a position effect or gene interaction (a position effect or gene interaction effect)effect)The Rh Ag on the red cell The Rh Ag on the red cell is normalis normal, but the , but the steric arrangement of the C Ag in relationship to steric arrangement of the C Ag in relationship to the D Ag appears to interfere with expression of the D Ag appears to interfere with expression of the D Ag.the D Ag.

This interference with D expression does not This interference with D expression does not occur when the C gene is inherited in the cis occur when the C gene is inherited in the cis (The location of two or more genes on the same (The location of two or more genes on the same chromosome of a homologius pair) .Family study chromosome of a homologius pair) .Family study can be distinguish which type of weakened D Ag can be distinguish which type of weakened D Ag is being demonstrated. is being demonstrated.

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D MosaicD Mosaic One or more parts of the One or more parts of the D Ag are missingD Ag are missing

Wiener and Unger postulated that the complete Wiener and Unger postulated that the complete D (Rho) Ag had four parts designated RhD (Rho) Ag had four parts designated RhAA, Rh, RhBB, , RhRhCC, Rh, RhDD , a lowercase superscript of , a lowercase superscript of a, b, c, and a, b, c, and dd is used to indicate when the corresponding is used to indicate when the corresponding portion (s) of the mosaic is missing.portion (s) of the mosaic is missing.

Example, RhExample, Rhbb means the B portion is absent. means the B portion is absent.

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Rh ReagentsRh Reagents

Coomb’s cells (C.C)Coomb’s cells (C.C)Control cellular used when Control cellular used when AHG test is negativeAHG test is negative..To confirm that washing has been adequate and To confirm that washing has been adequate and the anti-globulin reagent the anti-globulin reagent is reactiveis reactive

Control CellControl Cell

A group A group O Rh positive donor’sO Rh positive donor’s sample is mixed sample is mixed with with 1:10 dilution of reagent anti-D1:10 dilution of reagent anti-D and and allowed allowed to incubateto incubate..After After sensitization of RBCsensitization of RBC, they are washed with , they are washed with saline and suspended to a saline and suspended to a 50% RBC suspension50% RBC suspensionThese sensitized RBCs then used to confirm the These sensitized RBCs then used to confirm the anti-IgG activity of the AHG.anti-IgG activity of the AHG.

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Rh ReagentsRh Reagents

Coomb’s cells (C.C)Coomb’s cells (C.C)

They can be used to ensure that AHG test with neg They can be used to ensure that AHG test with neg results are not results are not false negfalse neg because of because of inactivation inactivation of of the the AHGAHG reagent. reagent.

When When AHG test is negAHG test is neg , they should be free , they should be free AHGAHG reagent in the test tubereagent in the test tube

When the When the CCCC are added, the free are added, the free AHGAHG in the test in the test should cause should cause agglutinationagglutination of the sensitized RBCs. of the sensitized RBCs.

This positive reaction is a mixed field in nature This positive reaction is a mixed field in nature because;because;

half of RBCs in the mixture lack half of RBCs in the mixture lack IgGIgG on their on their surface and are free cells surface and are free cells

22ndnd half of RBCs have half of RBCs have IgG IgG ((CCCC) and are agglutinated) and are agglutinated

Drmsaiem Rh Blood Group System DR. MOHAMMED H SAIEMALDAHR BB Faculty of Applied Medical Sciences MED...

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