Reducing the artifacts produced by impure antisera in immunoblots of low-molecular-mass proteins in urine.
Immunoblots of several urinary low-molecular-mass proteins can be very useful in investigations of pathological proteinuria. However, use of certain commercial antisera in such procedures leads to artifacts corresponding to nonspecific bands; e.g., immunoglobulins from nonimmunized rabbit serum may bind to human urinary proteins, and this binding is not inhibited by Triton X-100. We have developed a procedure to improve the specificity of detection of urinary low-Mr proteins separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, by immunoblotting with commercial antisera: we treat the protein blot with a mixture of mercaptoethanol and sodium dodecyl sulfate before incubation with the first antiserum. This allows direct use of commercial antisera without prior absorption of contaminating antibodies.
J M
JM
Perini
B
B
Dehon
T
T
Marianne
A
A
Klein
P
P
Roussel
D000510:Alpha-Globulins / Q000652:urine; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007106:Immune Sera; D007163:Immunosorbent Techniques; D008623:Mercaptoethanol / Q000494:pharmacology; D008722:Methods; D009113:Muramidase / Q000652:urine; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D011507:Proteinuria / Q000652:urine*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D001613:beta 2-Microglobulin / Q000652:urine
D000510:Alpha-Globulins; D007106:Immune Sera; C025500:alpha(2)-microglobulin; D001613:beta 2-Microglobulin; D012967:Sodium Dodecyl Sulfate; D011092:Polyethylene Glycols; D008623:Mercaptoethanol; D017830:Octoxynol; D009113:Muramidase