Rh BLOOD GROUP SYSTEMRh BLOOD GROUP SYSTEM

AHLS 311

HISTORYHISTORY

Ab in serum of mother of stillborn child; responsible for the death of fetus? (1939, Levine and Stetson)

Rb-derived Ab to Rhesus monkey RBCs reacts with 85% of human subjects; same Ab as reported by Levine? (1940, Landsteiner and Weiner)

Erythroblastosis fetalis (HDN) linked with Anti-

Rh (1941, Levine et al)

NOMENCLATURE: NOMENCLATURE: 4 VERSIONS4 VERSIONS

Fisher Race Suggested 3 sets of closely linked alleles (D

and d, C and c, E and e) Each gene (except d, which is an amorph)

causes production of an Ag Inherited from parents in linked fashion as

haplotypes See Tables 6-1 and 6-2

NOMENCLATURENOMENCLATURE

Weiner Multiple alleles at 1 complex locus 1 locus encodes for production of an

agglutinogen which has 3 factors (antigens or epitopes)

Abs can recognize single or multiple factors See Table 6-3

WEINER’S THEORYWEINER’S THEORY

WEINER & FISHER-RACE WEINER & FISHER-RACE TERMINOLOGYTERMINOLOGY

WEINER & FISHER-RACE WEINER & FISHER-RACE TERMINOLOGYTERMINOLOGY

1 ( C)

D C

2 ( E )

D c E

0 (neither C or E )

D c e

Z (both C & E )

D C E

‘( C)

d C e

‘’ ( E )

d cE

(neither C or E )

d c e

y (both C & E )

d C E

D = R

d = r

NOMENCLATURENOMENCLATURE

Rosenfield No genetic assumptions made Numerical system

If listed alone, the Ag is present (Rh:1 = D Ag)

If listed with a “-”, the Ag is not present (Rh:1, -2, 3 = DcE)

If not listed, the Ag status was not determined

Adapts well to computer entry

COMMON Rh TYPES BY COMMON Rh TYPES BY 3 NOMENCLATURES3 NOMENCLATURES

NOMENCLATURENOMENCLATURE

Internatl. Soc. of Blood Transfusion 6 digit number for each Ag specificity First 3 indicate the blood group, eg., 004 =

Rh Last 3 indicates the Ag specificity, eg.,

004001 = D Ag of Rh system For recording of phenotypes, the system

adopts the Rosenfield approach

Rh PHENOTYPINGRh PHENOTYPING Uses

Parentage testing Predicting hemolytic disease of the newborn

(HDN) Confirmation of Rh Ab specificity Locating compatible blood for recipients

with Rh Abs Protocol

Mix unknown RBCs with Rh antisera Take tubes through phases (IS,

heat/potentiator, AHG, CCC); record data Use published frequencies and subject

information to determine genotype

Rh GENOTYPING PHENOTYPING DATA Reactions with Anti-:

POSSIBLE GENOTYPES

D C E c e 1st CHOICE

2ND

CHOICE + + - - +

- - - + +

+ + - + +

+ + + + +

GENOTYPE GENOTYPE FREQUENCIESFREQUENCIES

Dce (R1) 0.42 dce (r) 0.37 DcE (R2) 0.14 Dce (R0) 0.04 dCe (r’) 0.02 dce (r”) 0.01 DCE (Rz) <0.01 dCE(ry) <0.01

The probability of 2 frequencies appearing together = the product of those 2 frequencies. For example, DCe/dce occurs with a frequency of 0.42 X 0.37 or 0.155.

Rh ANTIGENSRh ANTIGENS

Nonglycosylated proteins (A,B,H are CHOs) Transmembrane molecules D and CE are epitopes of proteins with 417 Aas

that traverse the membrane 12 X DNA sequences of D and CE differ by only 44

base pairs; CE, Ce, cd and cE are even more similar to D

Integral part of RBC membrane (Rhnull people have mild hemolytic anemia)

Density of Rh Ags on RBCs varies by phenotype (see Table 6-7)

MODEL OF Rh PROTEINMODEL OF Rh PROTEIN

D ANTIGEN VARIATIONSD ANTIGEN VARIATIONS Weak D

Some cells require addition of AHG (IDAT) to demonstrate agglutination with Anti-D

3 mechanisms causing weak D expression Genetic - inheritance of D genes which

result in lowered densities of D Ags on RBC membranes

C trans - position effect; the D gene is in trans to the C gene, eg., Dce/dCe (C and D Ag arrangement causes steric hindrance weakening D expression)

D mosaic - 1 or more parts of the D Ag is missing; may result in production of Anti-D

People with weak D are considered Rh+ and receive Rh+ blood (except mosaics)

D ANTIGEN VARIATIONSD ANTIGEN VARIATIONS

Enhanced D When c and D are in double doses, eg.,

cDe/cDe, (C has limiting effect on expression of D)

D-- or D .. represent partial locus deletions; usually seen in consanguinous situations

D TESTINGD TESTING Anti-D reagents

Saline-based - Low protein (fewer false positives); long incubation times; cannot convert to weak D testing

Protein-based - Faster, increased frequency of false positives; requires use of Rh control tube, converts to weak D testing

Chemically modified - “Relaxed” form of Anti-D in low protein medium; few false positives; saline control performed; converts to weak D testing

Blends of mAbs

D TESTINGD TESTING Protocol

Add Anti-D to “D” tube; Rh control to “C” tube Spin, read and record

If “D” is positive, cells are Rh positive If “D” is negative, continue testing

Add 22% albumin and incubate for 20” at 37oC Spin, read, and record Wash 3 X in saline Add AHG, spin, read, and record If “D” is positive after heat/albumin or AHG

cells are weak D positive; if negative, cells are Rh negative; “C” should always be negative

Add check cells to neg. tubes; spin, read & record

WEAK D Ag IN THE LABWEAK D Ag IN THE LAB Differences from normal D expression

Quantitative (inherited weak D or position effects)

Qualitative (mosaic D; could produce Anti-D) If cells are weak D, consider the person to be Rh +

Dw not given to D negative recipients D positives usually OK for Dw recipients Dw mothers do not receive RhoGAM

Donors and expectant mothers should be tested for weak D; transfusion recipiencts +/- for weak D testing (Dw people may receive D negative blood)

OTHER ALLELES AND OTHER ALLELES AND ANTIGENSANTIGENS

Weak C (Cw) Not allelic to C and c (C and Cw usually seen

together) 2% of whites; very rare in blacks Anti-Cw may be naturally occurring and

shows dosage f (ce)

When c & e are in cis, eg., dce/DCe Combination Ag Anti-f may be helpful in phenotyping

OTHER ALLELES AND OTHER ALLELES AND ANTIGENSANTIGENS

Ce When C and e in cis Compound Ag Ab helpful in phenotyping

G Always found with C-positive RBCs; usually

with D-positive cells Anti G appears to bind to D, C, and G

Many others

ALLELIC DELETIONSALLELIC DELETIONS No Cc and/or Ee epitopes

DC-, Dc-, D-E, D-- Enhanced or exalted D Ag expression

Rhnull (no Rh Ag expression at all) ---/--- (double bar rr) Or, because of independently inherited

suppressor genes If exposed to any Rh Ags, make Abs to

those and to Rh 29 (“pan” or “total” Rh) Causes a mild hemolytic anemia

Rhmod - weakened expression of all Rh Ags

Rh ANTIBODIESRh ANTIBODIES

Immune IgG Abs (IgG1 and IgG3 most important)

React optimally at 37oC or with AHG Order of immunogenicity:

D > c > E > C > e Do not bind complement (RBC destruction by

Rh Abs is extravascular)

Rh Abs: Rh Abs: CLINICAL SIGNIFICANCECLINICAL SIGNIFICANCE

Severe HDN Severe transfusion reactions