Development and Application of an Immunocapture PCR Diagnostic Assay Based on the Monoclonal Antibody for the Detection of Shigella

Document Type: Letter

Authors

1 Kunming University of Science and Technology

2 Kunming Biomed International

3 Faculty of Life Science and Technology, Kunming University of Science and Technology, Kunming, China

4 Research Center of Molecular Medicine of Yunnan Province, Faculty of Life Science and Technology, Kunming University of Science and Technology, 727 Jingming South Road, Kunming 650500, P.R. China.

10.30498/ijb.2020.127412.2244

Abstract

Background: Shigella is regarded as the most important human pathogenic microorganisms, infecting both humans and animals, and causing clinically severe diarrhea. The necessary procedure of Shigella enrichment before detection is quite time-consuming.
Objectives: Development of a sensitive, rapid, and specific method for Shigella detection.
Materials and Methods: The Shigella was used as an antigen to generate monoclonal antibodies (mAbs). mAbs were carried out on screening by indirect enzyme-linked immunosorbent (ELISA) and Western blot, and two mAbs were selected successfully. The mAb A3, with high affinity and specificity, was selected to develop the Immune magnetic beads (IMB) for the enrichment of Shigella. Meanwhile, a pair of IC-PCR primers were designed according to the ipaH gene. Then, the immunocapture PCR (IC-PCR) was developed based on the IMB and PCR.
Results: In this system, the time for the detection of Shigella was shortened to 70 min. Results demonstrated that the sensitivity of the IC-PCR was 9 CFU mL-1 in artificial milk. Besides, the accuracy of the IC-PCR was identified by 46 clinical samples collected from monkeys. The results of the IC-PCR were consistent with the serological and biochemical assay.
Conclusion: In conclusion, the IC-PCR described here can screen Shigella in milk samples and monkeys accurately, which could be used as a complement for classical detection methods.

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