Measurement of cytochrome f in thylakoids of higher plants using quantitative gel electrophoresis.

A quantitative method was developed to estimate the concentration of cytochrome (cyt) f in isolated thylakoids, using sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by staining with a heme-specific reagent containing 3,3',5,5'-tetramethylbenzidine and hydrogen peroxide. This densitometric technique was at least as sensitive as difference spectroscopy. Analysis of thylakoid preparations by densitometry of stained bands using cyt c as standard gave molar ratios of cyt/chlorophyll which were identical to ratios obtained by difference spectroscopy. Densitometric assays demonstrated that the molar ratio of cyt f/chlorophyll decreased during leaf aging in seven higher plants; however, there was a marked difference in the rate at which cyt f was lost from the leaves of different species.