Antibody Screening. Antibody Screening in Pre-transfusion Testing and Antenatal Screening

Transcription

1 Antiody Screening in Pre-trnsfusion Testing nd Antentl Screening

2 Antiody Screening in Pre-trnsfusion Testing nd Antentl Screening Q. Wht re nturlly occurring or expected ntiodies? Q. Wht re typicl or unexpected ntiodies? Q. How typicl ntiodies re formed nd which re cliniclly significnt ntiodies? Q. Wht re utontiodies? Q. Wht is pre trnsfusion testing? Q. Wht is Antiody Screening? Q. How Antiody Screening is done? Q. Wht is Antiody Identifiction? Q. Wht is the difference etween Crossmtch nd Antiody Screening? Q. Wht is Type nd Screen Policy? Q. Wht is the importnce of Antiody Screening in ntentl cses? Nturlly occurring or expected ntiodies. If n ABO ntigen is missing from n individul's red cell memrne, then it is EXPECTED tht the individul will produce n ntiody to tht ntigen. These hve lso een clled nturlly occurring ntiodies. Exmple: nti-a, nti-b, nti-ab. Helthy dults lwys hve nti-a nd/or nti- B ntiodies in their serum if they lck the corresponding ntigen on their red cells. Atypicl or Unexpected ntiodies. In ll lood group systems other thn ABO, if the ntigen is missing from the red cell, the individul is NOT expected to produce n ntiody ginst it, normlly. When these ntiodies re produced they re termed UNEXPECTED or ATYPICAL or ALLOANTIBODIES. The production of these ntiodies is result of n event, like lood trnsfusion or Pregnncy. These ntiodies re normlly of IgG clss. Exmple: nti-d, nti-c, nti Fy. Formtion of Atypicl or Allontiodies. Allontiodies re the ntiodies produced ginst foreign ntigen. These ntiodies cn e mde in response to trnsfusion of red cells or exposure to fetl red cells during pregnncy or delivery. These ntiodies re directed to non-self ntigen nd thus clled s llontiodies. There re now out 270 uthenticted lood group ntigens. Mny of these lood group ntigens fll into one of 26 lood group systems. Aprt from these 26 Blood group systems there re few more ntigens which re not ssigned to ny lood group system. Antiodies to ll these ntigens re not cple of cusing Hemolytic Trnsfusion Rection or HDFN. There re some ntiodies which re cliniclly significnt nd cple of cusing Hemolytic Trnsfusion Rection nd HDFN, such few ntiodies re listed elow. 1

3 Nme of Blood Group System Cliniclly Significnt Antiodies ABO A, B & AB Rh D, C, E, c, e Kell K Kidd Duffy Jk, Jk Fy, Fy MNS S, s (rrely M, N) Lewis Le (rrely) Autontiodies. Autontiodies re ntiodies, usully formed y disese process or mediction. These ntiodies re produced ginst person's own red cells. Pre-trnsfusion testing. The ojective of pre-trnsfusion testing is to ensure tht enough red cells nd components will survive when trnsfused, or in other words, to trnsfuse lood component to ptient tht will provide mximum enefit while cusing less hrm. In most lood nks, pre-trnsfusion testing involves (i) determining the ABO nd Rh types of ptient nd donor lood, (ii) screening ptient nd donor ser for RBC llontiodies, nd (iii) performing mjor crossmtch. Pre-trnsfusion testing cn ssure ABO comptiility etween donor nd ptient lood s well s detect most cliniclly significnt RBC llontiodies tht cn rect with donor's red cell ntigens. Antiody screening. Aprt from clericl checking, grouping nd typing of donor nd ptient lood, the serum or plsm of the ptient must e tested ginst pnel of group O regent red cells. Such regent red cells re selected ecuse they crry the lood group ntigens necessry for detecting the most importnt "cliniclly significnt" RBC llontiodies. This procedure is known s ntiody screening nd these regent red cells re known s screening cell pnels. The ntiody screening determines whether n ntiody to red cell ntigen hs een produced. Antiody screening is performed to detect ntiodies in: Ptients requiring lood trnsfusion Women who re pregnnt Ptients with suspected trnsfusion rections Blood nd plsm donor 2

4 Antiody screening cells re regent red cells tht provide comintion of ntigens other thn A nd B ntigens. These cells re tested with ptient's serum/plsm to determine whether n unexpected ntiody exists. Below is n exmple of n ntigrm for three cell screening pnel. An ntigrm lists the ntigens present in the red cell suspensions. A rection to either of the screening cells demonstrtes the presence of n typicl ntiody. Three cell pnel is lwys preferred over two cell pnel ecuse it provides n rr (Rh Negtive) cell nd homozygous cells for the Duffy nd Kidd lood groups. Most common, cliniclly significnt ntiodies rects with three cell pnel nd initil conclusions regrding the type of ntiody cn often e mde when screen is complete. G E L S Y S T E M ERYGEN AS Antigrm. Antiody Screening procedure. Screening cell pnel consists of red cells of three donors' leled s Cell I, Cell II nd Cell III. Ptient's serum is tested with these regent red cells in n IAT phse. Commercil red cell pnels re supplied in pproprite cell suspension which mkes them redy to use. In Mtrix Gel System totl procedure for ntiody screening is of 30 minutes. 50µl of TM regent red cells to e pipetted in to the leled microtues of Mtrix AHG (Cooms) test crd followed y 25µl ptient's serum or plsm. The crd is then incuted for 15 minutes t 37 C followed y centrifugtion. Results re then red nd interpreted on the ntigrm provided with the cell pnels. An utocontrol tests the ptient's serum with his or her own red cells. Testing n utocontrol routinely with the screen is optionl; most lood nkers prefer to perform DAT only if the screen is positive. The utocontrol nd DAT provide useful informtion in determining whether ptient's ntiody is directed ginst his or her red cells or trnsfused cells. Antiody identifiction. Antiody identifiction is performed fter the positive results of ntiody screening. In ntiody identifiction serum or plsm is tested ginst pnel of regent red cells. A pnel like the screening cells, consists of group O regent red cells tht hve een typed for most common ntigens specificities. Commercil cell pnels re ville with vriety of ntigen configurtions, which my include 10, 11, 15, 16 or 20 cells tht cn e thought of s extended ntiody screens. The use of utocontrol with the pnel is recommended, especilly if it is not routinely tested with the screening cell pnel. 3

5 Once testing is done results re recorded on ntigrm provided with the kit for ech phse of testing. The phse or rection temperture t which gglutintion ppers is n indiction tht the ntiody is IgG or IgM. IgM ntiodies typiclly rect t room temperture or on immedite spin. IgM ntiodies such s nti-le, -Le, -M, -N, -I nd P1 should e suspected if immedite spin rections re detected. IgG ntiodies rect t the ntigloulin phse. Rections t different phses indicte more thn one ntiody nd comintion of IgG nd IgM ntiodies. After recording the results on n ntigrm provided with the cell pnel the ntiody is identified y following steps. Ruling Out: Cells tht give negtive rection with ll tested phses cn e used to rule out the ntiodies. If the ntigen ntiody rection did not occur, this suggests tht the ntiody did not rect with the ntigen present on the pnel cell, nd respective ntigens cn e eliminted s possile ntiody. Antigens tht re heterozygous should not e cross out ecuse ntiody might hve een too wek to rect. This process is continued for ech negtive cell. Mtching the pttern: The next step in pnel interprettion is to look t the rections tht re positive nd mtch the pttern. When single ntiody is present, the pttern of rections oserved mtches with the other cells. Rule of three: In rule of three, t lest three ntigen positive red cells tht rect nd three ntigen negtive red cells tht do not rect should e oserved. Phenotyping the ptient: Individuls do not mke llontiodies to ntigens they possess. Another wy to confirm ntiody identifiction is to phenotype the ptient's red cells to ensure tht they re negtive for the ntigen corresponding to the identified ntiody. Crossmtch Vs Antiody Screening Antiody Screening is the most relile nd sensitive method of detecting llontiodies. Crossmtch is often less relile when compred with Antiody Screening, ecuse some ntiodies mnifest dosge effect. To explin the dosge effect we tke n exmple of Kidd lood group system hving two mjor ntigens Jk nd Jk. Plese refer to elow tle: Cell Jk Jk Remrks Cell I + 0 This cell is hving Homozygous expression of Jk Cell II 0 + This cell is hving Homozygous expression of Jk Cell III + + This cell is hving Heterozygous expression of Jk nd Jk In ove exmple, cell I crries the Homozygous expression of Jk nd results in higher expression of the Jk ntigen thn the cell III which crries the heterozygous expression of oth Jk nd Jk. While performing crossmtch, phenotype (ntigenic configurtion of red cells) of donor's red cells is not known, nd there re possiilities tht, ptient is hving nti- Jk nd donor red cell crries heterozygous expression of Jk. This my led to comptile crossmtch even in the presence of corresponding ntiody. For these resons, n ntigloulin crossmtch using donor cells is not the most effective wy of detecting serologicl incomptiility etween donor nd ptient. 4

6 A mjor crossmtch, involves testing of the ptient's serum with donor's RBCs, in IAT phse. In crossmtch we detect the presence of ntiodies in ptient's serum/plsm corresponding to the ntigens present on the donor's red cells. A Negtive or Comptile crossmtch shows tht ntiodies corresponding to the ntigens of donor's red cells re sent in ptient's serum/plsm. But comptile crossmtch does not signify tht there re no typicl ntiodies present in ptient's serum/plsm. Type nd Screen Policy. In some specil instnces, crossmtching of lood is excluded from pretrnsfusion testing ccording to policy clled "type nd screen." This policy stipultes tht lood does not hve to e crossmtched in dvnce for ptients undergoing surgicl procedures usully not requiring lood. The ptient's lood is, however, completely tested for ABO group, Rh type, nd RBC llontiodies nd then kept in storge y the trnsfusion service in cse it is needed for crossmtching. In most countries, typed nd screened ptient's lood cn e crossmtched y IS-XM (immedite spin cross mtch or sline crossmtch) nd mde ville in minutes, however in Indi lood nks following Type nd Screen policy performs Cooms crossmtch whenever lood unit is required for the ptient. Type nd Screen cn e very useful for the lood nks hving high Crossmtch: Trnsfusion (CT) rtio. CT rtio is the rtio of lood units crossmtched nd lood units trnsfused in hospitl. Higher CT rtio suggested tht numer of lood units crossmtched is higher thn the numer of lood units trnsfused. Before the introduction of this procedure, mny units of donor lood were crossmtched nd held in reserve for ptients who would proly not need it. At times this would cuse shortges of the lood supply nd unnecessry outdting of donor units. These fctors, long with the dded expense of crossmtching lood, cused the Type nd Screen (T&S) procedure to gin populrity. This procedure is used most frequently to screen pre-opertive or gynecologicl ptients whose risk of excessive lood loss is miniml. In cse of n emergency, where lood is needed for these ptients, IS-XM (sline crossmtched), ABO nd D comptile lood cn e relesed with 99.9% ssurnce of sfety, s long s the ptient hs no unexpected ntiodies. Antiody screening in Antentl cses. The ntigen tht most frequently induces immuniztion is D ut ny red cell ntigen present on fetl cells nd sent from the mother cn stimulte ntiody production. As these ntiodies re of IgG clss, they re cple of crossing plcent nd my cuse hemolysis of fetl red lood cells. HDFN is often clssified into three ctegories on the sis of the specificity of the custive IgG ntiody. In descending order of potentil severity they re: 1. D hemolytic disese cused y nti-d lone or less often in comintion with nti-c or nti-e. 2. Other hemolytic disese cused y ntiodies ginst other ntigens in the Rh system or ginst ntigens in other systems; nti-c nd nti-k re most often implicted. 3. ABO HDFN cused y nti-a,b in group O womn or y isolted nti-a or nti-b. 5

7 In order to detect these ntiodies the smples of ll pregnnt women should e tken erly in pregnncy idelly t weeks gesttion for ABO nd D typing nd for screening for the presence of red cell llontiodies. When n ntiody screen is positive further tests should e crried out to determine the ntiody specificity nd significnce. All pregnnt women whether D positive or D negtive, should hve further lood smple tken t 28 weeks gesttion for re-checking the ABO nd D group nd further screening for red cell llontiodies. D positive women re just s likely s D negtive women to form ntiodies other thn nti-d lte in pregnncy. When red cell ntiodies re detected, further testing of mternl lood should e undertken to determine the specificity, concentrtion, origin nd level of ntiody or ntiodies nd the likelihood of HDFN. Anti-D, nti-c nd nti- K re the ntiodies most often implicted in cusing hemolytic disese severe enough to wrrnt ntentl intervention. Biliogrphy 1. Bsics nd pplied concepts of Immunohemtology 2. BCSH Guidelines for pre-trnsfusion comptiility procedures in lood trnsfusion lortories. 3. BCSH Guidelines for lood grouping nd ntiody testing in pregnncy. 4. Guidelines for pre-trnsfusion testing; Austrlin nd New Zelnd Society of Blood Trnsfusion. 5. Current issues in trnsfusion medicine; Pretrnsfusion testing of red cells-current sttus 6. Mximum surgicl lood ordering schedule in tertiry trum center in northern Indi: A proposl. 7. th AABB, Technicl Mnul, 15 Edition. 6