Development Of Membrane Strip Tests For The Rapid Detection Of Cabaryl

Carbaryl is an N-methylcarbamate that is widely used as a broad-spectrum insecticide. In this paper, two competitive assays based on the principle of immunoassay using HRP and gold colloidal as tracers for the rapid on-site detection of carbaryl were developed.For immunoassay using HRP as tracer, there were two formats: flow-through and dipstick. Nylon~+ membrane from Millipore was used as solid support. The membrane was coated with anti-carbaryl antibody and a carbaryl-horseradish (HRP) conjugate was used as the labeled antigen for direct competitive assay of carbaryl. The optimal experimental condition was as follows: 1.0 μg of anti-carbaryl antibody coated on the membrane, the dilution of carbaryl-horseradish conjugate 1: 100000. Both tests had visual detection limits of 10 μg/Lfor carbaryl.For one-step gold-based immunoassay, there were also two formats: flow-through and lateral-flow. Nitro-cellulose membrane strip used as solid support was separately coated with goat anti-rabbit IgG (control line) and carbaryl hapten-OVA conjugate (test line). Anti-carbaryl antibody conjugated with gold colloidal was used as detectable tracer for carbaryl detection. The optimal experimental condition was as follows: 1.0 μg of carbaryl hapten-OVA coated on the membrane, incubation time between anti-carbaryl antibody labeled with gold particles and carbaryl was 5 minutes, the ratio of anti-carbaryl antibody labeled with gold particles and carbaryl for flow-through and lateral-flow were 1:3 and 1:2, respectively. The visual detection limits of carbaryl for flow-through and lateral-flow gold-based assays were 50μg/L and 100 μg/L.Chinese cabbage, rice, barley, apple and pear were used as real sample (negative sample detected by HPLC) for analysis. These food samples were spiked with carbaryl, mixed with 5 volume of methanol, and shaking by hand for 5 min. Matrix interference was eliminated by appropriate dilution of sample extracts with buffer. The membrane strip test can be used directly for the analysis of sample extracts. However, for HPLC analysis, the sample extracts should be purified with SPE. The results obtained by test strip were coherence with the results obtained by HPLC. A carbaryl-horseradish conjugate and anti-carbaryl antibody gold conjugate were placed at 4℃ and 37 ℃ for 7 days , membrane strips were tested for each day. Results showed that anti-carbaryl antibody gold conjugate was more stabilize than carbaryl-horseradish conjugate.These rapid tests can be useful as a quality control tool and can be used to semi-quantitative or qualitatively detect carbaryl in plentiful samples.