Insolubilization of protein antigens on polyacrylic plastic beads using poly-L-lysine.
A new model system for quantitation of immunofluorescence on single cells is described using poly-L-lysine (PLL) coated polyacrylic plastic beads of approximately cell sizes as carriers for protein antigen. By increasing PLL concentration on the beads increased amounts of 125I labeled antigen were attached to the particles. Excess binding sites of PLL could be completely blocked by unrelated proteins. After staining with FITC-conjugated antibodies and quantitative fluorescence measurements of individual beads using a microspectrofluorimeter, strong correlations were found between antibody and antigen concentration on the beads. Neither repeated washings with PBS nor storage of the beads for two months caused detectable shedding of antigen-antibody complexes. There was a strong linear correlation between fluorescence intensity and the volume of beads, but the correlation between surface area and fluorescence was nonlinear. The described procedure was shown to be a simple method for quantitative and stable coating of particles with proteins. It can be applied as a useful model system for quantitative immunofluorescence studies on intracellular antigens.
D000178:Acrylamides; D000941:Antigens; D005455:Fluorescent Antibody Technique / Q000379:methods*; D008239:Lysine; D008863:Microspheres; D008954:Models, Biological; D011108:Polymers; D011506:Proteins / Q000276:immunology*; D013050:Spectrometry, Fluorescence
D000178:Acrylamides; D000941:Antigens; D011108:Polymers; D011506:Proteins; D008239:Lysine