Detection of all proteins on western blots provides a visual image of the electrophoretic pattern, which helps identify specific antigens in a complex protein mixture. Western blot stains have a sensitivity comparable to gel stains.
Bio-Rad offers four stain options for total protein detection:
SYPRO Ruby protein blot stain — provides highly sensitive detection of proteins on PVDF or nitrocellulose membranes. The stain makes it possible to visualize sample proteins and molecular weight standards directly on the blot, eliminating the need to stain duplicate gels or guess at protein transfer efficiencies. After staining, target proteins can be detected by colorimetric or chemiluminescent immunostaining or analyzed by microsequencing or mass spectrometry with no interference from the stain.
Biotin-Blot total protein stain — optimized for nanogram level detection of total protein bound to nitrocellulose or Zeta-Probe nylon membranes. Membranes analyzed for total protein with the Biotin-Blot method can be conveniently compared with a duplicate membrane that has been probed with antibodies or other ligands. Antigenic proteins and immune complexes are easily correlated because direct comparisons of duplicate membranes eliminate the problem of evaluating gels that have undergone shrinkage and shape change. This detection method is based on the high affinity of avidin for biotin and the resulting stable avidin-biotin complex.
Coomassie Brilliant Blue R–250 dye — anionic dye used for staining gels and PVDF and nylon membranes. It is a rapid and inexpensive stain that can detect nanogram levels of protein. Since this dye can interfere with antibody binding sites, subsequent immunodetection of proteins is not recommended.
Colloidal gold total protein stain — stabilized gold solution optimized for rapid and sensitive identification of proteins bound to nitrocellulose membranes (Rohringer and Holden 1985). Protein bands stain dark red following incubation of the membrane with colloidal gold solution. The stained membrane yields a permanent record of the protein pattern for exact comparison to immunostained results. Colloidal gold total protein stain is provided ready to use.
Reference
Rohringer R and Holden DW (1985). Protein blotting: detection of proteins with colloidal gold, and of glycoproteins and lectins with biotin-conjugated and enzyme probes. Anal Biochem 144, 118–127.
