A sensitive and rapid method for the determination of protein by the resonance Rayleigh light-scattering technique with Pyrogallol Red

Abstract

The resonance Rayleigh light-scattering (RRLS) technique was used to develop a simple, sensitive and selective method for the determination of proteins. The method is based on the interaction between proteins and Pyrogallol Red (PR) in the pH range 3.6–4.2, which causes a substantial enhancement of the resonance scattering signal of PR in the wavelength range 300–450 nm with the maximum scattering peak located at 347 nm. With this method, 0.25–13 μg ml⁻¹ of bovine serum albumin (BSA), 0.25–10 μg ml⁻¹ of human serum albumin (HSA) and 0.25–13 μg ml⁻¹ of human immunoglobulin G (IgG) can be determined, and the detection limits, calculated as three times the standard deviation of nine blank measurements, for BSA, HAS and IgG were 51, 48 and 57 μg l⁻¹, respectively. Moreover, the method shows almost no protein-to-protein variability and is free from interference from many amino acids and metal ions. The method, with high sensitivity, selectivity and reproducibility, was satisfactorily applied to the determination of the total protein in human serum and saliva samples. Mechanism studies indicated that PR can bind to BSA depending mainly on electrostatic forces, and this interaction can encourage the J-aggregation of PR, which results in enhanced Rayleigh light-scattering in the PR–protein system.