Incorporation of bromophenol blue enhances visibility of polyacrylamide gels.

Vertical polyacrylamide gel electrophoresis (PAGE) is routinely used in molecular biology to separate macromolecules. DNA sequencing, singlestranded conformational polymorphism (SSCP) analysis and microsatellite typing all utilize the high resolution capabilities of polyacrylamide gels to separate molecules that differ in size by as little as one base pair. However, preparation of these gels is both tedious and time-consuming. One of the problems associated with polyacrylamide gels is that, before polymerization, they are highly neurotoxic (2). This is unfortunate, because the unpolymerized gel is a clear liquid and can be difficult to see if the gel leaks. Another problem is the occurrence of unwanted bubbles when the gel is poured (1). These bubbles, if ignored, can affect the normal migration of samples through the gel matrix after the gel has polymerized. This makes the identification of bubbles crucial when every lane of a gel is to be used. We have found that a common laboratory dye, bromophenol blue, greatly enhances the visibility of polyacrylamide gels. This enhanced visibility results in a gel solution that is brilliant blue and thus easy to see. The colored gel solution serves as an excellent warning to potential hazards, similar to the dye used in some commercially available radioisotopes. The color also causes any bubbles to contrast sharply with the gel when the gel is poured, thus allowing their removal before polymerization. The dye appears to have no effect on normal migration of samples through the gel. Since bromophenol blue migrates anodally during electrophoresis, the gel can be pre-run for a short while to remove any dye that may affect subsequent protocols (e.g., silver staining or chemiluminescense). A few minutes of pre-run time before the samples are loaded (which is a common practice used to bring gels up to operating temperature) is usually sufficient to clear the wells and top portions of the gel so that the dye used in the samples to be loaded is still visible, thus allowing assessment of inter-lane leakage. The dye front in the gel migrates at the same rate as the bromophenol blue in the samples; thus, the dye front itself can be used as an estimator of run-time. Alternatively, the gel can be pre-run so that the dye in the samples serves as an indicator of molecular size. We have not determined the effect of the dye on automated devices that analyze the gel automatically; however, once again a simple pre-run should alleviate any interference that may be present in the dye. We routinely incorporate 1–5 mg of bromophenol blue (Sigma Chemical, St. Louis, MO, USA) into a gel solution immediately prior to adding the catalysts necessary for polymerization (e.g., ammonium persulfate [Sigma Chemical] and N,N,N,N,-tetramethylenediamine [Bio-Rad, Hercules, CA, USA]). The powdered dye goes into solution very quickly. Our laboratory normally uses Sequagel (National Diagnostics, Atlanta, GA, USA), but we see no reason why this technique could not be used with straight polyacrylamide gels or with other commercially available solutions. This technique is safe, easy and cost-effective (less than $0.01/ gel), especially when compared to the cost of reactions that may be lost and/or unscorable due to interference with bubbles during electrophoresis.