Preparation, stability and commutability of candidate reference materials for lactate dehydrogenase (LDH).

Qi Guo, Zejian Zhu, Jing Wang, Wei Huang, Chao Zhang, Jie Zeng, Haijian Zhao, Tianqi Qi, Weiyan Zhou, Tianjiao Zhang, Chuanbao Zhang, Fei Xiao

Clinical biochemistry 2021 May

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Lactate dehydrogenase (LDH) is a key enzyme that functions as a marker of cell damage. Its activity can be measured by a variety of laboratory methods. To eliminate inter-method bias and enhance equivalence among different measurement procedures, LDH detection needs to be standardized. Optimized sequences coding for lactate dehydrogenase subunit A (LDH-A) and subunit B (LDH-B) were synthesized and cloned into the pRSFDuet vector, and then the constructed 6His-LDHA-pRSFDuet, 6His-LDHB-pRSFDuet, and 6His-LDHA-LDHB-pRSFDuet plasmids were transformed into Escherichia coli BL21 (DE3) for expression. The enzyme activity and specific activity of recombinant LDHs were detected. Electrophoresis of LDH isoenzymes was performed. The stability of recombinant LDHs and serum LDH was evaluated. Commutability of recombinant LDH-B was studied by the IFCC reference method and six routine methods. Three plasmids were constructed and three highly concentrated recombinant LDH isoenzymes were obtained. The specific activities of LDH-A, LDH-AB, and LDH-B were 18.08 U/mg, 21.74 U/mg, and 14.18 U/mg, respectively. There was a desirable linear correlation between the activities of recombinant LDH isoenzymes and their protein concentrations. Electrophoresis of LDH isoenzymes showed that the recombinant LDH-B corresponded to LDH1 and it demonstrated good stability at 4 °C and 25 °C for 5 weeks. LDH-B formulations in saline-bovine serum albumin solution and human serum matrix were commutable for six routine methods. Human recombinant LDH-B has great potential to become an improved and less expensive standard or reference material in external quality assessment for clinical LDH measurement. Copyright © 2021 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

Citation

Qi Guo, Zejian Zhu, Jing Wang, Wei Huang, Chao Zhang, Jie Zeng, Haijian Zhao, Tianqi Qi, Weiyan Zhou, Tianjiao Zhang, Chuanbao Zhang, Fei Xiao. Preparation, stability and commutability of candidate reference materials for lactate dehydrogenase (LDH). Clinical biochemistry. 2021 May;91:45-51