Key comparison study on protein quantification: purity-assessed recombinant protein contents in buffer solution using insulin analogue

Main text The key comparison CCQM-K151 protein quantification: Purity-assessed recombinant protein contents in buffer solution using Insulin analogue was coordinated by Korea Research Institute of Standards and Science (KRISS) under the auspices of the Protein Analysis Working Group (PAWG) of the Consultative Committee for Amount of Substance: Metrology in Chemistry and Biology (CCQM) CCQM. Nine National Metrology Institutes and Designated Institutes participated. Participants were required to assign the mass fraction of human insulin analogue (Ins) present as the main component in the comparison sample for CCQM-K151. This study provided the means for assessing measurement capabilities for determination of purity-evaluated recombinant proteins up to 10 kDa containing up to 3 disulfide cross-links in an aqueous calibration solution using amino acid based ID-LC-MS. Sample purity was confirmed by coordinating lab. Nine results were submitted. All results applied the LC-MS/MS method and isotope dilution mass spectrometry (ID-MS) method for quantification of Ins by amino acid analysis through acid hydrolysis. Among nine results, the Key Comparison Reference Value (KCRV) was assigned with six results. The assigned KCRV was the arithmetic mean of the six results, of 3440 mg/kg with a combined standard uncertainty of 13.5 mg/kg, the k-factor for the estimation of the expanded uncertainty of the KCRVs was chosen as k=2.57. Three participants, reported lower values than the other six, requested another set of test materials for additional experiments to investigate source of discrepancy in their results, and therefore were excluded from KCRV estimation in CCQM-K151. Three participants progressed substantial experiments with further investigation into their negative-biased factor, and actively proved their measurement capability. The degree of equivalence (with the KCRV) and its uncertainty was calculated for each result. Six of the participants were able to demonstrate the ability to quantitatively determine purity-assessed recombinant protein in buffer solution. To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database kcdb.bipm.org/. The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).