I am running an assay for activity of mitochondrial complex III. Most protocols use sodium azide / KCN as an inhibitor of complex IV to prevent re oxidation on cytochrome c. However, on addition of sodium azide, I am getting a lower rate of reaction than when running it without NaN3. Also, the rxn reading at time 0 is different from the blank reading at time 0 when sodium azide is added (both blank and rxn have sodium azide , the only difference is that the blank does not have mitochondria). But in absence of sodium azide, both rxn and blank have same readings at time 0. Could anyone provide any insights as to what the reason may be and if it may be better to run the assay without NaN3?
KCN you have to have in the reaction mix but I am not so sure about NaN3. As an example: http://www.ncbi.nlm.nih.gov/pubmed/22653162
Could you provide a link to the protocol you use.
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