Coomassie Brilliant Blue R-250 (CBBR-250) is the most frequently used dye stain in protein research. However, relatively poor sensitivity and long application durations are the major limitations of this stain. In the present study, we have investigated the effect of pre-stain methanol washing on the sensitivity of the conventional CBB staining method. Concentrations of methanol ranging from 5 to 30% were prepared in 5% acetic acid. Pre-stain washing of SDS-gels using acetic acid (5%) and methanol were assessed at various timings and temperatures. Our results demonstrate that pre-stain washing by methanol greatly improves the sensitivity of the CBB stain and facilitates the visualization of low molecular weight components present in a complex muscle protein, actomyosin. The outcome of this study also revealed duration of pre-stain washing and temperature as other factors that determine the sensitivity of CBB staining method. Therefore, it is suggested that pre-stain washing with methanol may improve the sensitivity of CBB stain in SDS-PA gels.
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