Housekeeping proteins: A preliminary study illustrating some limitations as useful references in protein expression studies

A primary objective of many protein expression studies is to define expression patterns that can distinguish between normal and diseased states, enabling a better understanding of molecular events associated with disease development and progression and ultimately potentially finding novel markers or therapeutic targets. Exploration and confirmation of many proteins is often done using Western blotting with normalization against housekeeping proteins, such as glyceraldehyde-3-phosphate dehydrogenase (GAPDH), beta-actin, or beta-tubulin, to correct for protein loading and factors, such as transfer efficiency. Increasingly, in studies examining gene transcript levels, it has been shown that some of the commonly used housekeeping genes may be unsuitable due to the influence of various physiological and pathological factors on their expression. This has not been examined to any great extent for proteins, however. This study examines the degree of variability of three commonly used housekeeping proteins (GAPDH, beta-actin, and beta-tubulin) together with class I beta-tubulin, with comparisons being made between a number of different established renal cancer cell lines, matched pairs of renal tumor and normal kidney lysates as well as nine different human tissues and highlights some of the problems encountered.