Galkin A. Biotechnology of obtaining and use of monoclonal antibodies to human IgM

The thesis presents the results of studies aimed for elaboration of biotechnological fundamentals of human anti-IgM MAb production and investigation. An original set of 21 novel clone hybridomas - producers of monoclonal antibodies to human IgM - has been obtained. Advanced study of biotechnological characteristics of the antibodies has been carried out: their specificity, affinity constant, and titer in culture medium have been determined. The epitope structure of human IgM molecule has been found to contain three basic epitope regions (ER): А, В, and С. ER А includes 1 predominant epitope А1, represented by the highest number of high affinity MAbs, and 2 adjacent epitopes А2 and А3 of medium affinity. ER В contains 2 adjacent epitopes В1 and В2 with medium affinity MAbs. ER С includes only 1 epitope with medium affinity MAbs. Epitope specificity of anti-IgM MAbs has been proven to depend on immunization scheme. The short-term immunization scheme results in mono-specificity of anti-ER-A antibodies, and the prolonged scheme causes the produced MAbs to be directed against antigenic determinants ER-B and ER-C. A possibility of using 2 MAb-based conjugates in test systems to diagnose cytomegalic inclusion disease and Epstein-Barr viral infection, as well as 3 MABs within a test kit immunosorbent for diagnostics of toxoplasmosis, rubella, urogenital chlamidiosis, and simplex herpes has been proven. A high sensitivity immunoenzyme kit for total IgM assay of analytical sensitivity 11 ng/ml has been elaborated. A well-reproducible and easy method of specific human IgM isolation using the produced anti-IgM MAb-based immunoaffine sorbent has been proposed.