I did a western blot detected using the supersignal west pico plus ECL reagent. However when imaged the blot background was dark grey and the bands white? Does anyone have an explanation as to why this has happened? I loaded my standard 10ug protein per well.
It may be that you have loaded too much material and in the band area it has depleted the ecl reagent completely in the time it takes to start measuring the signal. It may be that the problem will disappear with a lighter sample loading
Typically, when using chemiluminescence, when the signal saturates the band appears white. Ideally, dilutions of this antibody should be tested until an acceptable concentration is found.
Hi Stella,
Can you provide a copy of the image to have a look at? You may simply just need to invert the image to get the white background/dark bands you were expecting.
You can do this easily with our Phoretix 1D analysis software and automatically check for things like band saturation as mentioned in Vitor's reply: https://totallab.com/products/1d-gel-western-blot-analysis-software/
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