2026-11-3

On-Site Detection of Pasteurella multocida in Cattle: 15-Minute RPA Amplification with Lateral Flow Readout

A study in Irish Veterinary Journal reports an RPA-CRISPR/Cas12a workflow for detecting Pasteurella multocida in cattle — here is where EZassay's RPA kit fits in it.

Cover graphic: on-site detection of Pasteurella multocida in cattle — 15-minute RPA amplification

Bovine respiratory disease (BRD) remains a major challenge in cattle production. Identifying bacterial pathogens such as Pasteurella multocida often depends on laboratory-based methods, which can limit access to timely results in resource-limited settings.

A recent study by researchers at the College of Veterinary Medicine, Qingdao Agricultural University, demonstrates the potential of an RPA-CRISPR/Cas12a workflow for on-site pathogen detection. Targeting kmt1, a species-specific gene of P. multocida, the researchers developed an assay combining recombinase polymerase amplification (RPA), CRISPR/Cas12a detection, and lateral flow strip (LFS) readout.

RPA amplification was completed in 15 minutes at 37 °C. Following CRISPR/Cas12a detection, results could be visually interpreted using a lateral flow strip, without requiring a thermal cycler or fluorescence reader.

Detection workflow: target kmt1, a species-specific gene of P. multocida; DNA template from nasal swabs using a commercial extraction kit; RPA amplification at 37 °C for 15 minutes with the EZassay RPA kit; Cas12a with crRNA at 37 °C for 30 minutes; lateral flow strip read by eye after 30 minutes at room temperature, competitive format where a positive sample shows the C line only. Detection limit 10 copies/µL on plasmid standards, no cross-reactivity in the tested panel, 40 archived and 42 clinical bovine nasal swabs tested against conventional PCR and real-time qPCR respectively

Study Highlights

The study reported a detection limit of 10 copies/µL on plasmid standards, with no cross-reactivity against ten other tested bovine pathogens, including Mannheimia haemolytica, Histophilus somni, and bovine viral diarrhea virus.

In a blinded validation against conventional PCR, the assay showed complete agreement across 40 archived bovine nasal swab samples collected from cattle with clinical signs of BRD at commercial farms (17 positive, 23 negative).

In an additional cohort of 42 clinical nasal swab samples, the assay showed agreement with real-time qPCR across all 42 samples, with reference Ct values ranging from 16.0 to 35.0 (36 positive, 6 negative).

These data describe the complete detection workflow developed in the study, not the performance of any single reagent component within it.

Where EZassay's RPA Kit Fits in This Workflow

Integrated Product Portfolio

EZassay offers RPA amplification reagents and lateral flow strips as part of its broader molecular detection product portfolio, giving customers the convenience of sourcing both amplification and visual readout products from one supplier.

Integrated UDG Contamination Control

EZassay's contamination-control RPA kit incorporates a UDG-based strategy to help reduce the risk of aerosol carry-over contamination.

Published Use of EZassay RPA Kit

The study reports the use of EZassay's commercial RPA kit in a 20 µL reaction system according to the manufacturer's protocol. The Methods section states:

The RPA reaction was performed in a 20 µL reaction system using a commercial RPA kit (Shenzhen EZassay Biotechnology Co., Ltd., China) according to the manufacturer's protocol, with the integration of Uracil-DNA Glycosylase (UDG) system to prevent aerosol carry-over contamination. — Han X, et al. Rapid on-site detection of Pasteurella multocida in cattle using an LFS-based RPA-CRISPR/Cas12a assay. Irish Veterinary Journal. 2026. DOI: 10.1186/s13620-026-00360-4 · PMID 42806371

The Methods attribute the RPA kit to EZassay; the UDG system's supplier is not specified in the paper.

Three Practical Takeaways

  • 1.Application Potential. RPA-CRISPR/Cas12a offers a promising approach for rapid pathogen detection in veterinary applications.
  • 2.Contamination Control. UDG-based strategies are an important consideration in nucleic acid amplification workflows.
  • 3.Practical Readout. Lateral flow strips enable visual interpretation without specialized fluorescence readers.

EZassay RPA Product Line

  • ✔Efficient nucleic acid amplification
  • ✔Lyophilized format for improved storage and shipping flexibility
  • ✔Support for DNA amplification and RT-RPA workflows for RNA targets
  • ✔Reagents for fluorescence-based and lateral flow detection workflows
  • ✔Application support and assay customization
RPA isothermal amplification kits: liquid, lyophilized and UDG contamination-control formats, covering basic, lateral flow and fluorescent options, for DNA amplification and RT-RPA workflows for RNA targets
RPA core enzyme components: T4 UvsX, T4 UvsY, T4 Gene 32, Bsu DNA polymerase, endonuclease III and IV, RT-RPA reverse transcriptase, creatine kinase, UDG and UGI, with supporting reagents

Q&A

Whether you're at the primer screening stage or developing a POCT product for scale-up, our team can help identify suitable isothermal amplification reagents and workflows for your application.

EZassay provides a range of isothermal amplification reagents for basic, fluorescence-based, and lateral flow detection workflows, alongside Cas proteins and other tools for molecular detection research.

Our UDG-based contamination-control RPA kit helps reduce the risk of carry-over contamination, while our lyophilized RPA format offers greater flexibility for ambient-temperature storage and shipping. Both address practical challenges in developing POCT products for settings where cold-chain logistics may be difficult.

For application advice, technical references, or product selection support, contact our team at meg@ezassay.com.

Products are for research use only (RUO), not for clinical diagnosis.

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