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J Chromatogr B Analyt Technol Biomed Life Sci. 2019 Jul 25;1126-1127:121732. doi: 10.1016/j.jchromb.2019.121732. [Epub ahead of print]

Certification and stability assessment of recombinant human growth hormone as a certified reference material for protein quantification.

Author information

1
Center for Bioanalysis, Department of Chemical and Medical Metrology, Korea Research Institute of Standards and Science, 267 Gajeong-ro, Youseong-gu, Daejeon 34113, Republic of Korea; Department of Bio-Analytical Science, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 34113, Republic of Korea.
2
Center for Bioanalysis, Department of Chemical and Medical Metrology, Korea Research Institute of Standards and Science, 267 Gajeong-ro, Youseong-gu, Daejeon 34113, Republic of Korea; College of Pharmacy, Kyung Hee University, Seoul 02447, Republic of Korea.
3
Center for Bioanalysis, Department of Chemical and Medical Metrology, Korea Research Institute of Standards and Science, 267 Gajeong-ro, Youseong-gu, Daejeon 34113, Republic of Korea.
4
College of Pharmacy, Kyung Hee University, Seoul 02447, Republic of Korea.
5
Division of Medical and Biological Measurement, National Institute of Metrology, Beijing 100013, People's Republic of China.
6
Bio-Medical Standard Section, National Metrology Institute of Japan, National Institute of Advanced Industrial Science and Technology, 1-1-1 Umenozo, Tsukuba, Ibaraki 305-8563, Japan.
7
Center for Bioanalysis, Department of Chemical and Medical Metrology, Korea Research Institute of Standards and Science, 267 Gajeong-ro, Youseong-gu, Daejeon 34113, Republic of Korea; Department of Bio-Analytical Science, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 34113, Republic of Korea. Electronic address: jsjeong@kriss.re.kr.

Abstract

A certified reference material (CRM) for the quantification of protein, essential to manage quality control and quality assurance in protein-related works, has been developed. Amino acid analysis with conventional acid hydrolysis and isotope dilution HPLC-MS was used to establish an SI-traceable absolute protein quantification method using recombinant human growth hormone (hGH) as a model protein. The certification method was verified by comparative studies between 1) different methods of protein quantification based on microwave-assisted hydrolysis, and 2) different labs as part of the Asian Collaboration on Reference Material project with Japan, China, and Korea. Certification, evaluation of measurement uncertainty, homogeneity testing, and stability testing were carried out, after which the candidate CRM for hGH quantification was successfully certified with excellent agreement within the certified value in the two comparative studies. Although the quantification value of hGH by amino acid analysis showed good robustness in various conditions, results of intact protein analysis showed degradation profiles in temperatures higher than 4 °C. Consequently, storage and dissemination conditions should be set in accordance with stability tests. Based on the results, this method is believed to be suitable for accurate quantification of hGH. Additionally, it can also be used as a guide to preparation of CRM, and instructions for quality management of protein work for other similar proteins.

KEYWORDS:

Certified reference material; Human growth hormone; International comparison; Protein quantification; Stability assessment

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