We have further characterized the sensitivity and specificity of SYPRO Orange protein gel stain and SYPRO Red protein gel stain with native and 2-dimensional polyacrylamide gels and for staining gels prior to Western blot analysis. We found that nucleic acids are not stained by the SYPRO protein gel stains, in contrast to results obtained with commonly used silver staining techniques. Bacterial lipopolysaccharides also stain very weakly with SYPRO dyes in comparison to silver staining. Thus, gels containing whole cell lysates from either bacterial or mammalian cells can be analyzed more easily with the SYPRO stains than by silver staining. In this paper, we demonstrate that SYPRO stains can be used to monitor protein induction in bacterial overproducing strains and are effective stains for 2-dimensional gels. In addition, we developed protocols to detect proteins with these dyes in native gels. Finally, we found that staining proteins in transfer buffer prior to Western blotting does not affect the sensitivity of subsequent immunodetection.