RF detected by agglutination of tanned IgG-loaded erythrocytes is involved in negative idiotypic regulation of lymphocytes specific to autoimmunity-inducing antigens.

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The N. Blokhin National Cancer Research Center is one of the few Russian scientific institutions in which hybridoma technology of monoclonal antibody (mAb) production has been successfully established. Using this technology, several dozens of mAbs to various antigens of human leukocytes have been elaborated. These mAbs are widely used for immune status evaluation and for differential diagnostics of leukemias. Two mAbs were used to develop therapeutic drugs. Imuteran is a pharmaceutical form of mAb ICO-25 against a mucin-like antigen of human milk fat globules and proposed for treatment of epithelial cell-originating cancers (breast, intestinal, ovarian, lung cancer, etc.). ThePhase II clinical study of this agent is now nearly completed, and preliminary results suggest Imuteran to be a promising anticancer agent with tumor-stabilizing activity, but patients should be carefully monitored for signs of allergic reactions. mAb ICO-90 against the CD3 antigen of human T lymphocytes was used to develop the therapeutic agent Atemonate proposed for treatment of acute transplant rejection. At present, the Phase II clinical study of this agent is over, and the results confirm the drug safety and efficacy for this indication. The drug is being registered at the Ministry of Healthcare and Social Development, and transfer to serial production is expected shortly.

Fluorescent probes based on monoclonal antibodies (MAb) are widely used in scientific and clinical research in the field of oncology, hematology, immunology, epidemiology. Objective: to create of fluorescent probes based on the MAb and the fluorescent dye Alexa-488 for the analysis of cellular populations by flow cytometry. Materials and methods. MAb to B lymphocyte antigen (clone ICO-180), fluorescent dye Alexa-488 were used in the work. MAb was isolated from ascitic fluid by combined purification of the immunoglobulin fraction with caprylic acid and salting out with ammonium sulfate. Gel filtration on a PD-10 column was used to purify the conjugates (immunofluorescent probes, IFP), the concentration and labeling density of the IFP were determined spectrophotometrically. The determination of the working titer of the IFP was performed using the antibody titration method proposed by C.C. Stewart. Results. The optimal time of incubation of MAb with a fluorophore was experimentally determined. The optimal conditions for labeling MAb of the IСO series with the dye are: a carbonate buffer with pH 8,3, the concentration of antibodies in the reaction mixture is 1 mg/ml, molar ratio of active dye – 10–100 mmol per 1 mmol of protein, the incubation time is 90 minutes, the temperature is 18–25 °C. We obtained a panel of conjugates of MAb with Alexa-488, differing in their different labeling densities. Evaluation of the biological activity of the resulting conjugates was carried out on peripheral blood cells of donors in the concentration range of MAb 0,5–100 μg/ml. Conclusion. The optimal conditions for labeling MAb of the IСO series with the dye are: a carbonate buffer with pH 8,3, the concentration of antibodies in the reaction mixture is 1 mg/ml, the incubation time is 90 minutes, the temperature is 18–25 °C. The optimum density of labeling is in the range 5–13,5 M:M, the optimal concentration of antibodies is in the range of 5–25 μg/ml.

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