Suppose, one is doing enzyme assay of an enzyme of a steroidogenic pathway. Is it necessary to deplete the steroids from the tissue/ cell homogenate before administering substrates to check enzyme activity? Do people normally do charcoal depletion assays for this?
I am asking this because I thought that the exogenous labelled substrates added would compete with the substrates already present in the tissue and would not give the accurate maximal enzyme activity. Please suggest