Strategies to minimize variability and bias associated with manual pipetting in ligand binding assays to assure data quality of protein therapeutic quantification

Bioanalytical laboratories require accurate and precise wetting to assure reproducible and accurate results for reliable data. Two areas where wetting differences among analysts lead to poor reproducibility are long term stability testing and sample dilution. The purpose of this paper is to illustrate the problems with manual wetting, describe an automation strategy to mitigate risks associated with manual wetting, and provide recommendations on a control strategy that properly monitors samples requiring dilutions We determined differences among various manual wetting techniques by analysts within a laboratory To reduce variability in wetting, a flexible modular liquid handling script was created on the Hamilton Microlab Star (HMS) to perform sample dilution, pre-treatment and plate loading. The script is capable of handling variable dilution factors Additionally, two dilution controls were prepared and tested at concentrations of high and mid quality controls (QC) These same dilution controls were incorporated into both pre-study validation and in-study QCs to monitor dilution processing and assay performance. Variability of manual wetting among 11 analysts was more negatively biased with increasing dilution. Forward and reverse wetting delivering different volumes contributed to the discordance The dilutional bias with manual wetting was eliminated using the liquid handler Total error of dilution controls was less than 20% The in-study pass rate was 100% Application of liquid handlers minimizes the variability and bias clue to manual wetting differences among analysts The incorporation of dilution QCs serves a dual purpose to monitor the dilution process of the samples as well as the binding assay performance (C) 2010 Elsevier B.V All rights reserved