Experiment 11

GROUP 2 EXPERIMENT NO 11: D (Rh) TYPING – DIRECT, SLIDE METHOD

1. Characterize completely completely the the Rh antigens antigens (D,C,E,c,e) (D,C,E,c,e) in terms of their biochemical biochemical nature nature and serologic  property.  property. The final result of gene action in RC groups is the production of a biochemical structure! in the Rh system it is a nonglycosylated protein. This means that there are no carbohydrates attached to the  protein. The gene products of R"D and R"CE are remar#ably similar in that both encode for proteins composed of $1% amino acids that tra&erse the cell membrane 1' times and that their seuence differs  by only $$ base pairs. The gene products of  RHCE, RHCe,  RHce, and RHcE and  RHcE are e&en more similar. C and c differ from one another in four amino acid positions, and one amino acid differentiates E from e. nly nly smal smalll loop loopss of the the Rh prot protei eins ns are are e*po e*pose sed d on the the surfa surface ce of the the RC and and pro& pro&id idee the the conformational reuirements for the serologic differences bet+een the Rh blood types. n comparison +ith - and ell () blood groups, -1 cells possess appro*imately 1./ 0 1/ antigens, +hereas homozygous homozygous ell cells ha&e ///  sites. The greatest number of D antigen sites is on cells of the rare Rh phenotype D22. (D22 cells carry only D antigen and completely lac# Cc and Ee.) "o+e&er, "o+e&er, of the commonly encountered encountered Rh genotypes, genotypes,  R2R'  R2R' cells possess the largest number of D antigen sites. 3ummary4     

Rh antigens are non2glycosylated Rh antigens are transmembrane polypeptides and are an integral part of the RC membrane "ighly immunogenic4 5e*t to - and  D6c6E6C6e 7eighs appro*imately 1821%$ #D 7ell de&eloped in early fetal life

'. 9i&e a listing of the Rh antigens antigens in their different different nomenclatures, nomenclatures, and and their freuency of of occurrence

FREQUENCY D: 85% Caucasians, 92% Blacks, 99% Asians C: 68% Caucasians, 27% Blacks, 93% Asians E: 29% Caucasians, 22% Blacks, 39% Asians c: 80% Caucasians, 96% Blacks, 47% Asians e: 98% Caucasians, 98% Blacks, 96% Asians

Researcher4 :-5;dce DCe>DCe dce>dce DCe>DcE DcE>dce R'R'

DcE>DcE

Researcher4 C-3, Bary -ntonette :. Reference4 Bodern lood an#ing and Transfusion =ractices by "armening

$. 7hat are the causes of false positi&e and false negati&e reactions in Rh typing by the slide method

a.

mproper and inadeuate +ashing of the red cell suspension may cause pseudoagglutination due to the presence of serum macromolecules in the suspension. This should cause a positi&e control as +ell.  b. The presence of strong autoagglutinins in the patients or donors serum may cause agglutination. =roper +ashing and the control are designed to pre&ent or detect this problem. c. -ntibody coating of the red cell (positi&e D-T) can cause a false positi&e reaction, particularly in the +ea# D test. - D-T +ill detect this occurrence. d. - false negati&e reaction may be seen due to the bloc#ing phenomenon. This occurs most commonly in "DA5 due to anti2D. 3ince the red cells antigen sites are hea&ily coated +ith maternal antibody, they may not react +ith the antiserum. e. Aalse positi&e or negati&e reactions may occur in the Rh test due to many of the technical errors.. f. RCs that react +ith one manufacturerFs anti2D reagent but not +ith another may ha&e a partial D antigen. Researcher4 C-3, Bary -ntonette :. Reference4 cache42u#+p2content>uploads>'/11>/>'112Rh2 Typing.docHIcdJ$IhlJenIctJcln#IglJph 8. 7hat is the clinical! significance or importance of Rh typing Transfusion Reactions • Rh antigens are highly immunogenic. The D antigen is themost immunogenic antigen outside the - system. 7hen anti2D is detected, a careful medical history +ill re&eal RC e*posure through  pregnancy or transfusion of products containing RCs. Circulating antibody appears +ithin 1'/ days of  a primary e*posure and +ithin ' to % days after a secondary e*posure. Rh2mediated hemolytic transfusion reactions, +hethercaused by primary sensitization or  secondary immunization, usually result in e*tra&ascular destruction of immunoglobulin2 coated RCs. The transfusion recipient may ha&e an une*plained fe&er, a mild bilirubin ele&ation, and a decrease in hemoglobin and haptoglobin. The direct antihuman globulin test is usually positi&e, and the antibody screen may or may not demonstrate circulating antibody. 7hen the direct antiglobulin test indicates that the recipientFs RCs are coated +ith g9, elution studies may be helpful in defining the offending antibody specificity. f antibody is detected in either the serum or eluate, subseuent transfusions should lac# the implicated antigen. "emolytic Disease of the 5e+born ("D5) • "D5 is briefly described here because of the historic significanceof the disco&ery of the Rh system in elucidating its cause. -s stated pre&iously, anti2D +as disco&ered in a +oman after deli&ery of a stillborn fetus. The mother reuired transfusion. The fatherFs blood +as transfused, and the mother  subseuently e*perienced a se&ere hemolytic transfusion reaction. :e&ine and 3tetson1 postulated that the antibody causing the transfusion reaction also crossed the placenta and destroyed the RCs of the fetus, causing its death. The offending antibody +as subseuently identified as anti2D."D5 caused by Rh antibodies is often se&ere because the Rh antigens are +ell de&eloped on fetal cells, and Rh antibodies are primarily g9, +hich readily cross the placenta. -fter years of research, a method +as de&eloped to pre&entsusceptible (Rh/ D2negati&e) mothers from forming anti2D, thus pre&enting Rh/(D) "D5. Rh2immune globulin, a purified preparation of g9 anti2D, is gi&en to a D2negati&e +oman during pregnancy and follo+ing deli&ery of a D positi&e fetus. Researcher4 C-3,