Precautions for the cleanliness evaluation of reusable medical devices by alkaline extraction of residual proteins

Abstract

We reassessed the residual protein quantitative method, combining 0.2 M NaOH extraction and the Coomassie brilliant blue (CBB) assay for the cleanliness evaluation of reusable medical devices. The bovine serum albumin (BSA) calibration curve slopes using various protein assay kits were significantly reduced under alkaline conditions, except with the Bradford reagent. When evaluating the characteristics of four CBB reagents, the Coomassie plus Protein Assay Kit and Protein Quantification Kit-Rapid directly quantified a small amount of BSA even under alkaline conditions. Variation in the BSA recovery rate was observed with Pierce^TM 660 nm Protein Assay Reagent modified for micro assay and the Bradford reagent. In case of two assay kits other than CBB, contrary to the Sensolyte OPA Quantification Kit, the Micro BCA Assay Kit showed remarkable quantitativeness even under alkaline conditions. The recovery rate of the serum protein spiked on the stainless steel plate with alkaline extraction was comparable to the extraction with water or mammalian protein extraction reagent. Polyacrylamide gel electrophoresis showed that BSA was significantly decomposed to peptide fragments by hydrolysis after alkaline treatments. These results indicated that residual protein extraction with 0.2 M NaOH is not essential, and accurate quantification depends upon the proper selection of a protein assay kit and the pH and protein concentration of the test solution.