Bioanalytical Discovery & Development Sciences, Janssen Research & Development, Spring House, PA 19477, USA.
Discovery Technologies & Molecular Pharmacology, Janssen Research & Development, Spring House, PA 19477, USA.
Bioanalysis. 2022 Jul;14(14):985-1004. doi: 10.4155/bio-2022-0058. Epub 2022 Sep 6.
Industry-standard guidance on method development and validation of hybrid LC-MS/MS assays for protein biomarkers, particularly on evaluation of parallelism, is lacking. Using a protein endogenous to humans and mice as a model analyte, a quantitative hybrid LC-MS/MS workflow was developed using a surrogate matrix approach with a recombinant form of the protein as the calibrant. The developed workflow identified a surrogate matrix, established parallelism between the surrogate and authentic matrices and assessed parallelism between the recombinant and authentic forms of the protein. The final method was qualified using precision and accuracy with recovery assessments. The established workflow can be used in future bioanalytical studies to develop effective hybrid LC-MS/MS methods for endogenous protein biomarkers.
行业标准指南中缺乏有关用于蛋白质生物标志物的混合 LC-MS/MS 分析方法的开发和验证的信息，特别是关于平行性评估的信息。使用人类和小鼠内源性蛋白质作为模型分析物，使用替代基质方法和蛋白质的重组形式作为校准品开发了定量混合 LC-MS/MS 工作流程。所开发的工作流程确定了替代基质，建立了替代基质和真实基质之间的平行性，并评估了蛋白质的重组形式和真实形式之间的平行性。最后使用回收率评估对精密度和准确度进行了方法验证。所建立的工作流程可用于未来的生物分析研究中，以开发用于内源性蛋白质生物标志物的有效混合 LC-MS/MS 方法。
