In the spirit of the Halloween season, it’s time to let our skeletons out of the lab closet and share some of those freaky Western blots. You know – those scary ones that you tuck away in the back of your notebook hoping you never have to look at them again. The ugly splotched blots that always seem to be peeking out when your advisor walks by….
I will make you a deal: I will show you mine if you show me yours.
It was a dark and stormy night
It is so dark that you can’t even see your bands. In fact, you may as well be using pre-exposed film. Even the quickest dash and shortest exposure can’t help you escape this fiend. Some good washing and proper titration might clear the fog.
Too many spirits at the party
I was probing for a specific protein, right? If you have also seen this nice ladder of bands, you might suspect your secondary antibody as the culprit. Run a no primary antibody control and titrate the secondary antibody to chase those spirits away.
Someone is haunting you
Transfer artifacts can lead to ghostly images on your film. Air bubbles prevent contact between the gel and membrane and result in cleared areas on your film. Always press out bubbles when setting up the transfer stack.
A spirit trail
Dirt and particulates can lead to a blotchy spirit trail scattered across your blot. Keep your blots clean and make sure all buffers are fully dissolved.
The invisible blot
This is the trickiest of all blots – the invisible spirit that haunts your film. Nothing shows on your film and you can only mark the position of the molecular weight markers and hope that after a day of rest you might see a ghostly image. You know you put the right antibodies on there and followed the protocol, but without the proper positive control, no one will ever believe you.
Make sure you chase all of the spirits out of your blots! Read our WiKi and other blog posts for tips to lower your background and have successful blots.
Share your ugly, freaky Western blots with us in the comments below. We’d love to see your skeletons!
Photo courtesy of Shaun Dunmall.
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