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A novel biosensor-based microarray assay for the visualized detection of CYP2C19 ∗2, ∗3, ∗4 and ∗5 polymorphisms

  • Jian-Min Xiao , Jie Xiong , Guan-Yang Kang , Qiang Li , Zhong-Yong Jiang , Jiu-Hao Chen , Li-Ling Wang , Fu-De Yao and Jia-Wu Song EMAIL logo
Published/Copyright: October 2, 2014
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Clinical Chemistry and Laboratory Medicine (CCLM)
From the journal Clinical Chemistry and Laboratory Medicine (CCLM) Volume 53 Issue 2

Abstract

Background: A number of studies have indicated that the conversion of clopidogrel to its active metabolite is reduced in patients who carry the CYP2C19 *2, *3, *4 or *5 loss-of-function allele, resulting in decreased response of platelet to clopidogrel treatment and worse cardiovascular outcome. The aim of this study was to develop a novel biosensor-based microarray to visually detect CYP2C19 polymorphisms.

Methods: The target DNA was amplified from regions flanking the respective alleles using 5′-biotinylated reverse primer, and plasmids were prepared for the respective alleles. High stringency reversed hybridization, horseradish peroxidase-labeled streptavidin reaction, and color development, with multiple washes in different steps, were carried out and the results were recorded with an optical camera. The gene chips were tested for specificity, detection limit, intra- and inter-batch variations using the constructed plasmids. Finally, 88 clinical samples were assayed with this microarray as well as direct sequencing.

Results: The results could be seen with the naked eye. Concordance tests indicated that for alleles *2, *3, *4, and *5, the κ values between this assay and plasmids all reached 1.000. The detection limit was 5×102 cells/mL. Concordance test between direct sequencing and the microarray assay using 88 clinical samples gave rise to the κ value of 0.983, and p<0.01, indicating very high concordance.

Conclusions: This novel biosensor-based microarray assay can amplify the signal in situ so that it can be detected by simple instruments or even the naked eyes. It is promising for clinical application in hospital laboratories.

Keywords: biosensor; clopidogrel; CYP2C19; microarray; polymorphism
Corresponding author: Jia-Wu Song, PhD, Department of Gastroenterology, the Fifth Affiliated Hospital of Sun Yat-Sen University; Zhuhai Sinochips Bioscience Co. Ltd., 4F, Building #2, 24 Jinfeng Road, West, Zhuhai City, Guangdong 519085, P.R. China, Phone: +86 756 3318449, Fax: +86 756 3318499, E-mail:

Acknowledgments

We thank the patients who participated in this research. This work was supported by grant from the Key Science and Technology Plan Projects of Dongguan City (2011105102015) and grant from the Science and Technology Plan Projects of Guandong Province (2012B020700001).

Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

Financial support: None declared.

Employment or leadership: None declared.

Honorarium: None declared.

Competing interests: The funding organization(s) played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.

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Supplemental Material

The online version of this article (DOI: 10.1515/cclm-2014-0700) offers supplementary material, available to authorized users.

Received: 2014-7-7
Accepted: 2014-9-2
Published Online: 2014-10-2
Published in Print: 2015-2-1

©2015 by De Gruyter

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https://doi.org/10.1515/cclm-2014-0700
Keywords for this article
biosensor; clopidogrel; CYP2C19; microarray; polymorphism

Articles in the same Issue

  1. Frontmatter
  2. Editorials
  3. The new and the old of heparin-induced thrombocytopenia
  4. Biological variation – reliable data is essential
  5. Biological variation: back to basics
  6. Review
  7. Influence of educational, audit and feedback, system based, and incentive and penalty interventions to reduce laboratory test utilization: a systematic review
  8. Opinion Papers
  9. Recent guidelines and recommendations for laboratory assessment of the direct oral anticoagulants (DOACs): is there consensus?
  10. Meeting report: present state of molecular genetics in clinical laboratories. Report on the VII European Symposium on Clinical Laboratory and In Vitro Diagnostic Industry in Barcelona
  11. Genetics and Molecular Diagnostics
  12. Non-invasive prenatal diagnosis of monogenic disorders: an optimized protocol using MEMO qPCR with miniSTR as internal control
  13. A novel biosensor-based microarray assay for the visualized detection of CYP2C19 ∗2, ∗3, ∗4 and ∗5 polymorphisms
  14. General Clinical Chemistry and Laboratory Medicine
  15. Preanalytical errors: a preliminary approach to the point of view of primary health care givers
  16. Comparison of Improvacuter™ tubes with BD Vacutainer™ tubes for various hormones in the aspects of stability and influence of gel separators
  17. Use of quality indicators to compare point-of-care testing errors in a neonatal unit and errors in a STAT central laboratory
  18. Serological features of antibodies to protamine inducing thrombocytopenia and thrombosis
  19. Comparison of three different immunoassays in the diagnosis of heparin-induced thrombocytopenia
  20. How the direct oral anticoagulant apixaban affects hemostatic parameters. Results of a multicenter multiplatform study
  21. Auto-validation of complete blood counts in an outpatient’s regional laboratory
  22. Performance evaluation of the digital cell imaging analyzer DI-60 integrated into the fully automated Sysmex XN hematology analyzer system
  23. Circulating keratan sulfate as a marker of metabolic changes of cartilage proteoglycan in juvenile idiopathic arthritis; influence of growth factors as well as proteolytic and prooxidative agents on aggrecan alterations
  24. Reference Values and Biological Variations
  25. Biological variation database: structure and criteria used for generation and update
  26. Cardiovascular Diseases
  27. The tumor necrosis factor-α –238G/A and IL-6 –572G/C gene polymorphisms and the risk of idiopathic dilated cardiomyopathy: a meta-analysis of 25 studies including 9493 cases and 13,971 controls
  28. Diabetes
  29. Multicentre evaluation of the Premier Hb9210 HbA1c analyser
  30. Infectious Diseases
  31. Determination of quality control limits for serological infectious disease testing using historical data
  32. Corrigendum
  33. Enrichment and enumeration of circulating tumor cells by efficient depletion of leukocyte fractions
  34. Letters to the Editors
  35. Influence of age and gender on red blood cell distribution width
  36. Effect of exhaustive running exercise on red blood cell distribution width
  37. Clinically useful samples and reference change value
  38. Loss of retinol stability in patient samples
  39. Evaluation of a new thyroglobulin sensitive assay in patients with differentiated thyroid cancer
  40. Caution in a case of highly discrepant carbohydrate antigen 19-9 values in an apparently healthy patient taking spirulina: a case report
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