[Histonet] about stained tissue used for protein assay
I have just started working on rabbit heart tissue and I am developing assays to study proteins through western blots. These rabbit tissues are stained with Evans blue and Triphenyl tetrazolium chloride (TTC)to study regional ishcemia. The problem I have is that when I try to perform a protein quantitation using a Bradford or BCA assay, these dyes interfere with the absorption readings.
I request you to kindly inform me what method can be used to correct the error in the absorption caused by the dyes. The amount of dye that these tissues can take up is not known and hence I am not able to apply a particular value to correct the error. I have tried various protein qauntification methods but at all these wavelengths there is some absorbance by these dyes.
It would be of great help if you could let me know the details about the protein quantification method that would be best to follow. Or atleast how to perform a correction for the background values given by these dyes.
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