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Updated: Jun 5, 2026
Development of a Lateral Flow Immunochromatographic Strip for Rapid and Quantitative Detection of Small Molecule Compounds
Published on: November 13, 2021
Development of an immunochromatographic strip for simple detection of penicillin-binding protein 2'
Hidehito Matsui1, Hideaki Hanaki, Megumi Inoue
1Laboratory for Antimicrobial Agents, Kitasato University, 5-9-1 Shirokane, Minato-Ku, Tokyo 108-8641, Japan.
Abstract:
Infections with methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-resistant coagulase-negative Staphylococcus (MR-CNS) are a serious problem in hospitals because these bacteria produce penicillin-binding protein 2' (PBP2' or PBP2a), which shows low affinity to β-lactam antibiotics. Furthermore, the bacteria show resistance to a variety of antibiotics. Identification of these pathogens has been carried out mainly by the oxacillin susceptibility test, which takes several days to produce a reliable result. We developed a simple immunochromatographic test that enabled the detection of PBP2' within about 20 min. Anti-PBP2' monoclonal antibodies were produced by a hybridoma of recombinant PBP2' (rPBP2')-immunized mouse spleen cells and myeloma cells. The monoclonal antibodies reacted only with PBP2' of whole-cell extracts and showed no detectable cross-reactivity with extracts from other bacterial species tested so far. One of the monoclonal antibodies was conjugated with gold colloid particles, which react with PBP2', and another antibody was immobilized on a nitrocellulose membrane, which captures the PBP2'-gold colloid particle complex on a nitrocellulose strip. This strip was able to detect 1.0 ng of rPBP2' or 2.8 × 10(5) to 1.7 × 10(7) CFU of MRSA cells. The cross-reactivity test using 15 bacterial species and a Candida albicans strain showed no detectable false-positive results. The accuracy of this method in the detection of MRSA and MR-CNS appeared to be 100%, compared with the results obtained by PCR amplification of the PBP2' gene, mecA. This newly developed immunochromatographic test can be used for simple and accurate detection of PBP2'-producing cells in clinical laboratories.
Insights
A new immunochromatographic test rapidly detects penicillin-binding protein 2
Area of Science:
- Microbiology
- Immunology
- Medical Diagnostics
Background:
- Methicillin-resistant Staphylococcus aureus (MRSA) and coagulase-negative Staphylococcus (MR-CNS) infections pose significant hospital risks due to their resistance to beta-lactam antibiotics.
- This resistance is mediated by penicillin-binding protein 2' (PBP2'), which has a low affinity for these drugs.
- Current identification methods, like oxacillin susceptibility testing, are time-consuming, delaying appropriate patient treatment.
Purpose of the Study:
- To develop a rapid and accurate diagnostic test for PBP2'-producing bacteria.
- To enable quick identification of MRSA and MR-CNS in clinical settings.
- To overcome the limitations of traditional antibiotic susceptibility testing.
Main Methods:
- Production of monoclonal antibodies against recombinant PBP2' (rPBP2') using hybridoma technology.
- Development of an immunochromatographic strip assay utilizing gold colloid-conjugated antibodies and immobilized antibodies.
- Testing the assay's sensitivity, specificity, and accuracy against bacterial cultures and PCR methods.
Main Results:
- The immunochromatographic test detected PBP2' within approximately 20 minutes.
- The assay demonstrated high specificity, with no cross-reactivity against 15 other bacterial species and Candida albicans.
- 100% accuracy was achieved in detecting MRSA and MR-CNS compared to PCR confirmation of the mecA gene.
Conclusions:
- A novel, rapid immunochromatographic test for PBP2' detection has been successfully developed.
- This test offers a simple, accurate, and fast alternative for identifying PBP2'-producing bacteria in clinical laboratories.
- The assay can significantly improve the timely diagnosis and management of MRSA and MR-CNS infections.
