Anti-bacterial activity mediated by human platelets.

Platelet-mediated antibacterial activity against gram-negative microorganisms has been investigated. Data show that human washed platelets possess the ability to partially inhibit growth of Salmonella typhi Ty-2 (a smooth strain) but not of two rough strains of Salmonella minnesota (R345-Rb and R595-Re). On the other hand, no antibacterial activity was noted in the presence of the gram-positive Staphylococcus aureus. This activity is dependent on the length of incubation of platelets with bacteria and apparently is not mediated by platelet soluble factors. Finally, lysis of platelets with Triton X-100 following their incubation with bacteria gives rise to the same degree of antibacterial activity observed in the untreated samples. This implies that the described platelet function does not rely on the possible trapping of bacteria in platelet aggregates.

Production and characterization of monoclonal antibodies to anti-human chorionic somatomammotropin by immunization with two free synthetic peptides.

The peptides corresponding to the fragments 135-140 and 166-174 of human chorionic somatomammotropin (hCS) were synthesized, and used to raise monoclonal antibodies to the native hCS molecule. The synthetic peptides were injected into BALB/c mice in the free form, i.e. not conjugated to a carrier, and the spleens were fused with Sp2/01Ag8 myeloma line to produce monoclonal antibodies. The antibodies produced belonged to the IgM and IgG classes and, once purified by affinity chromatography on hCS-Sepharose, they were covalently coupled to macroporous polystyrene beads and characterized by competitive radioimmunoassay. Their affinity constants were determined by elaborating the radioimmunoassay data by nonlinear regression analysis and they were found to range from 10(5) to 10(6) M-1. The evaluation of the affinity constant of the antibodies produced is always important as a measure of the immunogenicity of an antigen, particularly when synthetic peptides are used as immunogens.

Identification of a mixture of two different monoclonal antibodies by nonlinear regression analysis after solid phase radioimmunoassay with labeled antigen.

The results of competitive solid phase radioimmunoassay with labeled antigen, when analyzed by nonlinear regression, may reveal the presence of two different monoclonal antibodies, occasionally produced in a hybridoma culture. In fact, a hybridoma-derived antihuman chorionic somatomammotropin antibody prepared in our laboratories showed an unsatisfactory curve fitting when the experimental data were elaborated on the basis of a monoclonal model. The antibody was subjected to recloning, and gave two separate homogeneous monoclonal antibodies with different affinity constants.

Antigenic structure of human chorionic somatomammotropin: four epitopes revealed by monoclonal antibodies.

The characterization of epitope specificity of a panel of 15 monoclonal antibodies against human chorionic somatomammotropin (hCS), previously prepared in our laboratory, allowed us to distinguish 4 antigenic determinants on the hCS molecule. Experiments were carried out with competition and sandwich assays. The results allowed us to divide our MAbs into 4 groups, recognizing 4 different epitopes, and to select 4 MAbs each distinguishing a different epitope in order to pursue further studies on the antigenic structure of hCS.