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

Background: Shigella is among the most important human pathogenic microorganisms, infecting both humans and nonhuman and causing clinically severe diarrhea. Shigella must be enriched before detection, which is time-consuming. Objectives: To develop a sensitive, rapid, and specific method for Shigella detection. Materials and Methods: Shigella was used as an antigen to generate monoclonal antibodies (mAbs). mAbs were screened via indirect enzyme-linked immunosorbent assay (ELISA) and western blot. and two mAbs were selected. The mAb A3 showed high affinity and specificity and was used to develop immune magnetic beads (IMBs) for Shigella enrichment. An immunocapture (IC)-PCR primer was designed from the ipaH gene, and IC-PCR was developed based on the IMBs and PCR. Results: This system shortened the Shigella detection time to 70 min. The sensitivity of the IC-PCR was 9 colony-forming units.mL(-1) in artificial milk. The accuracy of the IC-PCR was confirmed using 46 clinical samples collected from monkeys. The IC-PCR results were consistent with the serological and biochemical assays. Conclusion: The IC-PCR described herein accurately detected Shigella from milk samples, monkeys and can thus be used to complement classical detection methods.