Im quite new to cell culture and have had some inaccuracies while harvesting my cells and therefore counting them using a CASY counter.
I reconstituted in what I thought to be 50,000 cells/ul, but my western blot shows major inconsistencies.
I was wondering if there is any way to normalise my loading using coommasie stained gels and Fiji or an alternative?
I would use BCA but my samples are in urea buffer and my resources are limited during this internship.
Any help would be super appreciated! Thank you!
hi
you can load minimum volume of cell lyste in each well for WB .then get the signal with beta-actin or GAPDH antibody so extract the cell lysate concentration through compare band intensity in software ( such as imageJ) .
https://lukemiller.org/index.php/2010/11/analyzing-gels-and-western-blots-with-image-j/
However, due to the fact that the exact concentration of the samples is not known, the accuracy of this method will not be high.
Note: you can set up with one sample and then developed for all samples .
good luck
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