MicroELISA assays of anti-HLA activity and isotype of human monoclonal antibodies.
Both monoclonal human antibodies to HLA-DR antigens and supernatants from oligoclonal B-cell lines can be conveniently screened for activity by microELISA assays which use 1/10 of the volume of reagents used in conventional ELISA assays. Target cells are fixed to the bases of wells in Terasaki plates, 5 microliters volumes of supernatants incubated in these wells, and target bound antibody detected by peroxidase-conjugated anti-immunoglobulin followed by the substrate ABTS(2,2'-azino-di-(3-ethylbenzthiazoline sulphonate). The plates are read on a micro EIA reader. Supernatants can also be assayed for immunoglobulin content and isotype in Terasaki plates by coating the wells with isotype-specific anti-immunoglobulin, adding test supernatant and developing with appropriate peroxidase-conjugated anti-immunoglobulin sera and ABTS. When assaying for immunoglobulin content, the plates can be read either with a reader or by eye. Advantages and modifications of these procedures are discussed. There are no apparent practically important disadvantages to these procedures as compared with more conventional ELISA assays.
Hancock
R J
RJ
U.K. Transplant Service, Bristol, England.
Yendle
J E
JE
Bradley
B A
BA
D000911:Antibodies, Monoclonal / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007132:Immunoglobulin Isotypes / Q000032:analysis*; D008832:Microchemistry
D000911:Antibodies, Monoclonal; D007132:Immunoglobulin Isotypes