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Absolute quantitation of human serum cystatin C: candidate reference method by 15N-labeled recombinant protein isotope dilution UPLC-MS/MS

  • Qiaoxuan Zhang , Min Zhan , Xiongqiang Peng , Xing Jin , Jun Yan , Pengwei Zhang , Junhua Zhuang , Liqiao Han ORCID logo EMAIL logo and Xianzhang Huang EMAIL logo
Published/Copyright: September 27, 2024
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Clinical Chemistry and Laboratory Medicine (CCLM)
From the journal Clinical Chemistry and Laboratory Medicine (CCLM) Volume 63 Issue 4

Abstract

Objectives

Serum cystatin C (CysC) is a reliable and ideal endogenous marker for accurately assessing early changes in glomerular filtration rate (GFR), surpassing the limitations of creatinine-based estimated GFR. To improve the precision of GFR calculation, the development of strategies for accurately measuring serum CysC is crucial.

Methods

In this study, the full-length CysC pure product and fully recombinant 15N-labeled CysC internal standard were subjected to protein cleavage. Subsequently, an LC-MS/MS method was developed for the absolute quantification of serum CysC. The traceability of the method was assigned calibrator using the amino acid reference measurement procedure (RMP). It involved calibrating the instrument using an amino acid reference material with known amino acid concentrations for calibration and comparison purposes.

Results

The total imprecision of the method was determined to be ≤8.2 %, and a lower functional limit of quantification (LLoQ) was achieved. The recoveries ranged from 97.36 to 103.26 %. The relative bias between this candidate RMP for measurement of ERM-DA471-IFCC and the target value was 1.74 %. The linearity response was observed within the concentration range of 0.21–10.13 mg/L, with a high R2 value of 0.999. The results obtained using our method was consistent with those obtained using other certified RMPs.

Conclusions

With the establishment of this highly selective and accurate serum CysC measurement method, it is now possible to assess the correlation between immunoassay results of serum CysC and the intended target when discrepancies are suspected in the clinical setting.

Keywords: reference measurement procedure; cystatin C; LC-MS/MS; standardization; 15N-labeled intact protein
Corresponding authors: Liqiao Han and Xianzhang Huang, Department of Laboratory Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine (Guangdong Provincial Hospital of Chinese Medicine), No. 111, Dade Road, Yuexiu District, Guangzhou 510120, Guangdong Province, P.R. China, E-mail: (L. Han); (X. Huang).
Qiaoxuan Zhang and Min Zhan contributed equally to this work.

Funding source: National Key Research and Development Program of China

Award Identifier / Grant number: 2022YFC3602300

Award Identifier / Grant number: 2022YFC3602301

  1. Research ethics: The collection of leftover serum samples was approved by the Ethics Committee of the Second Affiliated Hospital of Guangzhou University of Chinese Medicine (No. ZE2022-331-01).

  2. Informed consent: Because leftover samples involved in this study was anonymous, the ethics committee allowed the exemption of informed consent.

  3. Author contributions: The authors have accepted responsibility for the entire content of this manuscript and approved its submission.

  4. Use of Large Language Models, AI and Machine Learning Tools: None declared.

  5. Competing interests: The authors state no conflict of interest.

  6. Research funding: This research was supported by the National Key Research and Development Program of China (No. 2022YFC3602300 and No. 2022YFC3602301), “Young Talents Program” of Guangdong Academy of Traditional Chinese Medicine (No. SZ2022QN09), the Natural Science Foundation of Guangdong Province (No. 2021A1515220099 and 2023A1515220044), Guangzhou Science and Technology Plan Projects (No. 202201011506); the Research Fund for Bajian Talents of Guangdong Provincial Hospital of Chinese Medicine (No. BJ2022KY04); the Specific Research Fund for TCM Science and Technology of Guangdong Provincial Hospital of Chinese Medicine (No. YN2022QN16); the State Key Laboratory of Dampness Syndrome of Chinese Medicine (No. SZ2021ZZ30, SZ2021ZZ3003).

  7. Data availability: The raw data can be obtained on request from the corresponding author.

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Supplementary Material

This article contains supplementary material (https://doi.org/10.1515/cclm-2024-0300).

Received: 2024-03-05
Accepted: 2024-09-18
Published Online: 2024-09-27
Published in Print: 2025-03-26

© 2024 Walter de Gruyter GmbH, Berlin/Boston

Articles in the same Issue

  1. Frontmatter
  2. Editorial
  3. Beyond test results: the strategic importance of metadata for the integration of AI in laboratory medicine
  4. Reviews
  5. Reference, calibration and referral laboratories – a look at current European provisions and beyond
  6. How has the external quality assessment/proficiency testing of semen analysis been developed in the past 34 years: a review
  7. Opinion Papers
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  9. A comprehensive survey of artificial intelligence adoption in European laboratory medicine: current utilization and prospects
  10. Guidelines and Recommendations
  11. Guidelines for the correct use of the nomenclature of biochemical indices of bone status: a position statement of the Joint IOF Working Group and IFCC Committee on Bone Metabolism
  12. Candidate Reference Measurement Procedures and Materials
  13. Absolute quantitation of human serum cystatin C: candidate reference method by 15N-labeled recombinant protein isotope dilution UPLC-MS/MS
  14. General Clinical Chemistry and Laboratory Medicine
  15. Performance evaluation of the introduction of full sample traceability system within the specimen collection process
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  17. Comparison of capillary finger stick and venous blood sampling for 34 routine chemistry analytes: potential for in hospital and remote blood sampling
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https://doi.org/10.1515/cclm-2024-0300
Keywords for this article
reference measurement procedure; cystatin C; LC-MS/MS; standardization; 15N-labeled intact protein

Articles in the same Issue

  1. Frontmatter
  2. Editorial
  3. Beyond test results: the strategic importance of metadata for the integration of AI in laboratory medicine
  4. Reviews
  5. Reference, calibration and referral laboratories – a look at current European provisions and beyond
  6. How has the external quality assessment/proficiency testing of semen analysis been developed in the past 34 years: a review
  7. Opinion Papers
  8. Data flow in clinical laboratories: could metadata and peridata bridge the gap to new AI-based applications?
  9. A comprehensive survey of artificial intelligence adoption in European laboratory medicine: current utilization and prospects
  10. Guidelines and Recommendations
  11. Guidelines for the correct use of the nomenclature of biochemical indices of bone status: a position statement of the Joint IOF Working Group and IFCC Committee on Bone Metabolism
  12. Candidate Reference Measurement Procedures and Materials
  13. Absolute quantitation of human serum cystatin C: candidate reference method by 15N-labeled recombinant protein isotope dilution UPLC-MS/MS
  14. General Clinical Chemistry and Laboratory Medicine
  15. Performance evaluation of the introduction of full sample traceability system within the specimen collection process
  16. Pre-analytical stability of haematinics, lactate dehydrogenase and phosphate in whole blood at room temperature up to 24 h, and refrigerated serum stability of lactate dehydrogenase, folate and vitamin B12 up to 72 h using the CRESS checklist
  17. Comparison of capillary finger stick and venous blood sampling for 34 routine chemistry analytes: potential for in hospital and remote blood sampling
  18. Performance evaluation of enzymatic total bile acid (TBA) routine assays: systematic comparison of five fifth-generation TBA cycling methods and their individual bile acid recovery from HPLC-MS/MS reference
  19. Clinical performance of a new lateral flow immunoassay for xylazine detection
  20. Evaluation of revised UK-NEQAS CSF-xanthochromia method for subarachnoid hemorrhage: outcome data provide evidence for clinical value
  21. Strategies to verify equimolar peptide release in mass spectrometry-based protein quantification exemplified for apolipoprotein(a)
  22. Evaluation of the clinical performance of anti-mutated citrullinated vimentin antibody and 14-3-3 eta testing in rheumatoid arthritis
  23. Diagnostic performance of specific biomarkers for interstitial lung disease: a single center study
  24. Reference Values and Biological Variations
  25. Neonatal reference intervals for serum steroid hormone concentrations measured by LC-MS/MS
  26. Paediatric reference intervals for haematology parameters analysed on Sysmex XN-9000: a comparison of methods in the framework of indirect sampling
  27. Cardiovascular Diseases
  28. Analytical characteristics and performance of a new hs-cTnI method: a multicenter-study
  29. Diabetes
  30. Use of labile HbA1c as a screening tool to minimize clinical misinterpration of HbA1c
  31. Letters to the Editor
  32. Current trends and future projections in the clinical laboratory test market: implications for resource management and strategic planning
  33. Particulate matter in water: an overlooked source of preanalytical error producing erroneous chemistry test results
  34. “Activation” of macro-AST by pyridoxal-5-phosphate in the assay for aspartate aminotransferase
  35. The correlation of albumin with total protein concentrations in cerebrospinal fluid across three automated analysers – relevance to the diagnosis of subarachnoid haemorrhage in clinical chemistry practice
  36. Adult reference intervals for serum thyroid‐stimulating hormone using Abbott Alinity i measuring system
  37. Cell population data in venous thrombo-embolism and erysipelas: a potential diagnostic tool?
  38. Diagnostic performances and cut-off verification of blood pTau 217 on the Lumipulse platform for amyloid deposition in Alzheimer’s disease
  39. The first case of Teclistamab interference with serum electrophoresis and immunofixation
  40. Congress Abstracts
  41. Annual meeting of the Royal Belgian Society of Laboratory Medicine (RBSLM): “A Neurological Journey: Brain Teasers for Laboratory Medicine”
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