Spectrofluorometric and microfluorometric quantitation of antibacterial antibodies, using indirect immune...

The IgG-fraction of rabbit hyperimune serum raised towards smooth Salmonella typhimurium 395 MS (parent strain) and rough 395 MR10 (Rd-mutant) was used in indirect immunofluorescence with bacteria in suspension or adsorbed to glass microscope slides. In the first case, IgG-mediated fluorescence was determined with a standard cuvette-type spectrofluorometer; in the second case, in epifluorescence mode with a microscope fluorometer. In both assays the IgG solution could be diluted to about 0.1 micrograms per ml protein to allow detection of fluorescent anti-IgG antibodies in the second step. When determined by the spectrofluorometric method, the IgG antibody titres against S. enteritidis in two patients sera were 1/320 and 1/1280, respectively, which was compared to antibody titres determined with ELISA and MrPAH. Either mode of fluorescence measurement appears to give an objective but arbitrary value of the indirect immuno-fluorescence from bacteria and allows detection of antibodies directed towards whole bacteria, i.e. all antigenic determinants exposed. This may be of importance when following infectious processes. Furthermore, no extraction and binding of bacterial envelope components to solid supports is necessary.