Assays for Determination of Protein Concentration

Abstract

Biochemical analysis of proteins relies on accurate quantitation of protein concentration. This unit describes how to perform commonly used protein assays, e.g., Lowry, Bradford, BCA, and UV spectroscopic protein assays. The primary focus of the unit is assay selection, emphasizing sample and buffer compatibility. Protein assay standard curves and data processing fundamentals are discussed in detail. This unit also details high-throughput adaptations of the commonly used protein assays, and also contains a protocol for BCA assay of total protein in SDS-PAGE sample buffer that is used for equal loading of SDS-PAGE gels, which is reliable, inexpensive, and quick.

An analysis of critical factors for quantitative immunoblotting

“…Such estimates also do not account for variations in initial seeding density and final lysate volumes, which will affect the observed protein abundance. Consequently, immunoblot samples are typically prepared according to total cellular protein (18, 19), assuming that the average protein content per cell is constant across the different conditions.…”

Microcalorimetric assays for measuring cell growth and metabolic activity: Methodology and applications

“…optical methods such as spectrophotometry and microscopy are widely used to assess the growth of cell culture [6]. In addition, biochemical assays allowing the quantification of a cell component such as protein [14], sugar [15], DNA [16] or ATP [17] are also often used to estimate the number of cells in a culture. Finally in some context, measuring the impedance has been shown to be an accurate method to estimate cell number [13,[18][19][20].…”

Preparation and in vivo anti-tumor evaluation of human papillomavirus E7 adjuvanted with Montanide ISA 266 as a vaccine candidate

“…Finally, the purified proteins were dialyzed against phosphate-buffer saline (PBS; dialysis membrane 12 kDa cut-off) and stored at -70°C. The concentrations of the proteins were determined by Bradford assay [19].…”