Flow cytometry (FCM) analysis of live antibody-coated spermatozoa subjected to immunofluorescence staining (FCM test) is considered an objective method for the quantitative detection of antisperm antibodies (ASA). But the cross-linking of cell surface antigen (Ag) with bivalent antibodies and/or antigen-antibody (Ag-Ab) complexes with second antibodies may induce the reorganization of surface components (patching and capping) and result in their shedding from the sperm surface. The present study estimates the relationship between aggregation of Ag-Ab complexes on the sperm surface and the results of indirect FCM test. Swim-up spermatozoa of normozoospermic men were incubated with ASA-positive sera from infertile patients and with second antibodies fluorescein isothiocyanate (FITC)-labelled goat anti-human IgG polyclonal antiserum under different conditions and then analysed by FCM and fluorescence microscopy. It was shown that low temperature, cytochalasin B, excess or lack of the primary and/or secondary antibodies and sperm fixation by paraformaldehyde may inhibit aggregation and shedding of Ag-Ab complexes and dramatically increase ASA quantity determined on the sperm surface. However, inhibition of aggregation on the live sperm surface was observed only in a minority of ASA-positive samples and was poorly reproducible using semen of different donors. A high probability of Ag-Ab complex shedding from the sperm surface during experimental manipulation limits the use of indirect FCM test for quantitative ASA determination.

The cytotoxic activity of natural killer cells and the intensity of conjugate formation are studied in vitro in the natural cytotoxicity reaction against 3H-uridine-labeled human erythromyeloleukotic cells K-562 in the presence of fibronectin, T-globulin, and fibronectin/T-globulin combination. It is demonstrated that fibronectin does not change natural cytotoxicity, T-globulin increases the activity of human natural killer cells, and the fibronectin -T-globulin combination increases both the intensity of conjugate formation and the cytotoxic activity of natural killer cells. Key Words: natural killer cells; fibronectin; regulationA broad spectrum of interactions of human natural killer cells (NK) with different target cells (TC) is strongly influenced by the high degree of expression of adhesion molecules (LFA-1 and LFA-3) [12] and integrins (VLA-4 and VLA-5) [7,16] by them. Expression of these molecules on the NK surface correlates with the intensity of effector:target (E:T) conjugate formation [12]. These molecular complexes mediate not only the interaction of NK with the pericellular matrix components and other cells but also the adhesion to fibronectin (FN) [16,18] which is synthesized and expressed by NK [7,15] as a factor significant in the realization of the cytotoxic potential of the effectors of natural cytotoxicity (NCT) [15].The diversity of interactions between FN and different blood serum components implies that the Laboratory of Immunochemistry, N. F. Gamaleya Institute of Epidemiology and Microbiology, Russian Academy of Medical Sciences, Moscow effects of this protein in the organism are realized not by native but rather by modified molecules with considerably altered structural and functional characteristics due to partial catabolic cleavage involving R-proteins which exhibit affinity for FN [1], or resulting from the interaction of FN with aggregated IgG molecules which bind to both immobilized and dissolved FN [14]. Since serum IgG occur as complexes with proteins and protein components specifically bound to the C-terminal end of Fab-fragments [2], the above-mentioned interactions, including the formation of so-called natural antibodies during complexation of IgG with the products of partial catabolic cleavage of R-proteins [3], can change the delicate mechanisms of FN binding to the lymphocyte surface and the dynamics of cell-cell interactions realized via adhesion at the early stages, specifically, the reactions of antibody-dependent cellular cytotoxicity and lymphokme production [16].

It is demonstrated that in the presence of human recombinant T-interferon and Creactive protein the activity of natural killers increases 9-43%, whereas the number of effector:target cell conjugates formed at the stage of recognizing and binding of target cells decreases 14-80%. There is a weak positive correlation (p =0.35) between the activity of natural killers and the number of effector:target cell conjugates,

Antispermal antibodies were detected by flow cytofluorimetry and mixed antiglobulin reaction (MAR). The results of the two methods significantly correlated regarding selxun IgG antispermal antibodies under conditions ruling out capacitation of spermatozoa. There were no significant differences ha the content of antispermal antibodies in the sera of fertile, pregnant women, women with endometriosis and adhesions, or ial the peritoneal fluid of fertile and sterile women. In women with endometriosis and adhesions the level of antispermal antibodies in the peritoneal fluid was significantly higher than in the serum. The use of flow cytofluorimetry markedly improved the detection of antisperlnal antibodies in sterile women. Key Words: flow cytofluorimetry; antispermal antibodies; serumDespite numerous studies of the role of immune factors in sterility, the data on the contribution of antispermal antibodies (ASAB) to reproductive disorders in women are contradictory. These contradictions are due to the absence of a universal objective method for ASAB detection. Mixed antiglobulin reaction (MAR) is recommended by the WHO for identification of ASAB. However, this test is subjective and requires a labor-consuming microscopic analysis.Recently, flow cytofluorimetry (FCFM) was adapted for detecting ASAB in the ejaculate, male sera, and cervical mucus; this method possesses advantages over other methods of ASAB detection [1,[4][5][6]. The aim of this study was to explore the possibility of using FCFM for detecting ASAB in female serum and peritoneal fluid.