Author/Editor		Alscher, Maja
Title		Priprava monoklonskih protiteles ter reagentov, primernih za določanje antigenov krvnoskupinskega sistema AB0
Type		monografija
Place		Ljubljana
Publisher		Fakulteta za kemijo in kemijsko tehnologijo
Publication year		1995
Volume		str. 92
Language		slo
Abstract		Most of the reagents for phenotyping red cells available recently, were obtained from the plasma of people with suitable natural or immune abtibodies. However the use of polyclonal reagents with a changeable quality have became more and more questionable, especially when monoclonal antibodies (MAbs), produced by hybridoma technology, were introduced in blood group serology. In order to replace polyclonal reagents, produced at Blood Transfusion Centre of Slovenia (BTC) in Ljubljana with monoclonal ones, we selected approppriate antigens for the immunization of Balb/c mice. Spleen cells from such animals were fused with either P3-X63-Ag8.653 or P3-NS1/1-Ag4-1 mouse myeloma cell lines. Supernatants produced by hybridoma cultures were screened for specific antibody production by ELISA test, while selected hybridoma cell lines were chosen by performing agglutination tests, using mainly eritrocytes from newborn children. Promising cell lines were cultivated and stabilized several times by cloning. The final chosen clones were grown to maximal cell density in DMEM supplemented with 13% fetal calf serum (FCS). The nature of these antibodies was determined by specificity, potency and avidity tests. All of the produced MAbs against ABO antigens are of IgM type. Three series of test reagents were prepared out of seven clones of anti-A, anti-B or anti-A,B specifiy by cereful confectioning of supernatants. Stability tests were also performed. Finaly, reagents of the third series were chosen as the final diagnostic reagents, which were used in all routine procedures by testing a umber of samples in different laboratories of BTC in Ljubljana. The final anti-A and anti-B reagent are prepared with MAb C9/1,A5 and G1/2,B4, respectively, produced after immunizing mice with substance A from ovaorian cyst fluid and saliva of B secretor, while anti-A,B reagent is a blend of these anti-A and anti-B. Abstract truncated at 2000 characters.
Descriptors		ABO BLOOD-GROUP SYSTEM ANTIBODIES, MONOCLONAL BLOOD GROUPING AND CROSSMATCHING IMMUNIZATION, PASSIVE HYBRIDOMAS ANTIBODY SPECIFICITY INDICATORS AND REAGENTS REAGENT KITS, DIAGNOSTIC MICE