Lateral flow immunoassays (LFAs) are widely used for on-site testing; however, their use for the rapid detection of plant viruses in the field is often limited by inconvenient sample preparation. Here, we present a new sampling method and a simplified dipstick LFA format for the detection and monitoring of cowpea chlorotic mottle virus (CCMV) as a model plant pathogen. The assay employs a monoclonal mouse antibody for capture and a poly-clonal rabbit antibody conjugated to 80 nm gold nanoparticles for detection. Conventional sample and conjugate pads are omitted, allowing the test strips to be dipped directly into wells containing plant extract and antibody–gold conjugate. No plastic casing was required, which could lead to a reduction in waste. It was shown that CCMV concentrations as low as 3.5 µg/L or 350 pg per sample could be reliably detected in 15 min. Specificity tests confirmed that other plant viruses, cowpea mosaic virus (CPMV) and tobacco mosaic virus (TMV), did not produce false-positive results. In addition, we describe a new method for on-site sampling using a manual punch and a syringe equipped with a frit. This step combines grinding the sample, extraction, filtration, and reconstitution and mixing of the antibody-gold conjugate, enabling the analysis of punched leaf disks without laboratory equipment. When applied to CCMV-infected cowpea plants, the assay revealed systemic infection before visual symptoms became apparent. This work demonstrates that simplified LFAs combined with innovative sampling techniques can provide sensitive, specific, and rapid diagnostics for crop monitoring and support early intervention strategies in agriculture.

Lateral flow immunoassays (LFA) are widely used for decentralized testing, but their application for in-field plant virus diagnostics is often limited by tedious sample preparation. Here, we present a simplified dipstick LFA for the detection and monitoring of cowpea chlorotic mottle virus (CCMV) as a model plant pathogen. The assay employes a monoclonal mouse antibody for capture and a polyclonal rabbit antibody conjugated to 80-nm gold nanoparticles for detection. Conventional sample and conjugate pads are omitted, allowing the test strips to be dipped directly into wells containing plant extract and antibody-gold conjugate. In addition, no plastic casing was necessary, which significantly reduces waste. It was shown that CCMV concentrations as low as 4 µg/L or 400 pg per sample could be reliably detected in 15 minutes. Specificity tests confirmed that other plant viruses, cowpea mosaic virus (CPMV) and tobacco mosaic virus (TMV), did not produce false positive results. Furthermore, we describe a field-compatible sampling procedure using a manual punch and a disposable syringe. This step combines sample grinding, extraction, and conjugate reconstitution within the syringe frit, enabling the analysis of punched leaf discs without laboratory equipment. When applied to CCMV-infected cowpea plants, the assay revealed systemic infection before visual symptoms became apparent. This work demonstrates that simplified LFAs combined with innovative sampling techniques can provide sensitive, specific, and rapid diagnostics for crop monitoring and support early intervention strategies in agriculture.

Gene delivery lies at the heart of many approaches for treating a host of different diseases. Promising candidates for the delivery of genetic material are polycationic vectors; however, managing toxicity arising from adverse interactions with the lipid bilayer remains a challenge. In this work, photo-iniferter reversible addition-fragmentation chain-transfer PI-RAFT polymerization was used to synthesize statistical copolymers of N-vinyl formamide (NVF) and N-vinyl pyrrolidone (NVP). Subsequent selective hydrolysis of NVF was used to introduce polyvinyl amine (PVAm) repeats. The resulting library of polymers with varying charge densities and molar masses was probed for biocompatibility with erythrocytes and MDA-MB-468 cells, revealing substantially reduced cytotoxicity compared with linear polyethylene imine (lPEI) and Lipofectamine 2000. Using an ethidium bromide (EtBr) replacement assay PVAm copolymers were shown to replace EtBr at low N/P-ratios. The transfection conditions were optimized in terms of the N/P-ratio and polyplex concentration by a Renilla luciferase reporter assay. This revealed 30-fold less cytotoxicity, a much wider viable concentration range, and a twofold greater transfection efficiency for the PVAm copolymer compared to lPEI. This study provides insights into the PI-RAFT copolymerization of the less activated monomers NVF and NVP and highlights the potential of polyvinyl amine copolymers resulting from selective hydrolysis for the transfection of genetic material compared with lPEI.