Forbidden Technique: Bootleg Coomassie Blue

Out of Coomassie Blue 2 Hours Before Lab Meeting? Probably Not.

Coomassie blue, or SimplyBlue Safe Stain as we use in my lab, is a nonspecific protein dye used to allow visualization of SDS-PAGE bands. If you need some in a hurry there’s a quick and easy substitute that you probably have on hand: Bradford reagent. The normally red or rust colored Bradford reagent is the same exact dye as SimplyBlue, Coomassie Brilliant Blue G-250, differing only in solution pH.

Forbidden Technique: Bootleg Coomassie

To make bootleg SimplyBlue (probably not Safe Stain), aliquot around 40 mL of Bradford reagent and slowly add 3M NaOH (around 5 mL) until it becomes “brilliant blue”. Incubate this dye with your gel on a shaker for an hour or more and your bands will appear as normal, however you may notice some minor frothing if shaken vigorously. Proof below:

Figure 1. Lane 1: Dual protein standards. Lane 2-4: dCas9 (~169kDa) eluate from my most recent purification. These two gels are actually 1 gel cut in half and the only difference between them was the stain, SimplyBlue or Bradford pH 8. Gels were stained for 1 hour and are unwashed.

Do keep in mind that Bradford reagent is usually going to be more expensive than the bulk dye purchased for SDS-PAGE staining, but hey if you need results fast then price becomes subjectively irrelevant very quickly.

Additional note: sodium ions are noted in the SimplyBlue Safe Stain protocol to slow the staining process and thus overshooting with the pHing may be detrimental to your data. Take it easy, cowboy, you’re already going off-label so you may as well do it carefully!

I hope this trick brings you good fortune and even better results; the day I discovered this forbidden work around I had to present protein purification data in less than 2 hours and the panic brain blast truly came through.

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out /  Change )

Google photo

You are commenting using your Google account. Log Out /  Change )

Twitter picture

You are commenting using your Twitter account. Log Out /  Change )

Facebook photo

You are commenting using your Facebook account. Log Out /  Change )

Connecting to %s

%d bloggers like this: