To explore the feasibility of applying nanobodies in colloidal gold immunochromatography assay (GICA) for small-molecule harmful substances, this study systematically investigated the effects of colloidal gold labeling parameters, colloidal gold working buffer and sample pretreatment methods on GICA using the pesticide procymidone as a model. The results showed that the size of colloidal gold particles, labeling pH and the amount of antibody used were the key factors to ensure the stability of the nanobody-gold-labeled probes. Under the optimal conditions, the limit of detection, halfmaximal inhibitory concentration (IC₅₀) and visual detection limit of GICA for procymidone were 0.44, 6.29 and 200 μg/L, respectively. The sensitivity was 5.1-fold higher than that of conventional monoclonal antibody-based GICA. The quick, easy, cheap, effective, rugged and safe (QuEChERS) procedure was used for the pretreatment of Chinese leek, cucumber and tomato samples. Based on the excellent organic solvent tolerance of nanobodies, the organic solvent blow-drying procedure was omitted. The average recoveries of the spiked samples ranged from 80.1% to 109.6%, and the results for the actual samples were consistent with those of gas chromatography-mass spectrometry (GC-MS). This study showed that nanobodies are a potential alternative to conventional antibodies for immunoassays for the rapid detection of small molecules. The proposed nanobody-based GICA can be used for the rapid screening of procymidone in vegetables with the advantages of high sensitivity, high accuracy, rapidity and simplicity.

A matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) method was developed for the rapid detection of the emetic toxin cereulide in cooked rice and dairy products. The splitting pattern of cereulide standard was analyzed, meanwhile, effects of matrix type, spotting methods, the type and dosage of matrix solvent, and laser intensity on the MS signal intensity of cereulide were investigated. Methodological validation and application to actual samples were conducted. As a result, α-cyano-4-hydroxycinnamic acid (HCCA) was chosen as the matrix and dispersed in a 1:1 acetonitrile-water mixture containing 0.1% trifluoroacetic acid, and the matrix and the samples were sequentially spotted. In MALDI-TOF MS screening of foods using positive ion reflectron and linear modes at 70% laser intensity, the target ion peaks of ([M + Na]⁺ and [M + K]⁺) in cereulide were detected with stable signals, high intensity and good response repeatability. The results of methodological validation showed that good linear relationship was observed between the overlapping areas of [M + Na]⁺ and [M + K]⁺ peaks and cereulide concentration in the range from 5 to 100 ng/mL, with a correlation coefficients (r) greater than 0.99. The limits of detection (LOD) of [M + Na]⁺ and [M + K]⁺ were 3.0 and 5.0 ng/g, respectively; the recoveries of spiked rice and milk samples were ranged from 73.3% to 118.2%, with relative standard deviations (RSDs) of 0.3% to 10.9% (n = 6). This method was characterized by rapidity, high accuracy, good sensitivity, and strong anti-interference ability, and was suitable for the detection of cereulide in cooked rice and dairy products without using any internal standard.