Housekeeping Protein (HKP) Normalization Protocol v1

Housekeeping proteins (HKPs) are routinely used as loading controls for Western blot normalization. Accurate normalization requires stable expression of the HKP across all experimental conditions and treatments. Because HKP normalization relies on a single indicator of sample loading, variation in HKP expression leads to inconsistent estimation of sample loading and introduces experimental error that may alter data analysis. This protocol describes how to use a housekeeping protein for Western blot normalization and quantitative analysis. Note: This protocol is intended for use with near-infrared fluorescent Western blots.