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Ochratoxin A (OTA) is a common food contaminant that threatens consumers' safety and health. A sensitive and selective biotin-streptavidin-amplified enzyme-linked immunosorbent assay (BA-ELISA) for OTA using a nanobody-AviTag fusion protein (Nb-AviTag) was developed in this study. The prokaryotic expression vector Nb28-AviTag-pAC6 for Nb-AviTag was constructed, followed by transformation to the AVB101 cells for antibody expression and in vivo biotinylation. The purified Nb28-AviTag was used to establish the BA-ELISA and the procedures for this Nb-AviTag-based BA-ELISA were optimized. The Nb-AviTag-based BA-ELISA exhibited the half maximal inhibitory concentration (IC) of 0.14 ng mL and the limit of detection (LOD = IC) of 0.028 ng mL for OTA basing on the optimized experiment parameters. The assay sensitivity was improved 4.6 times and 4.3 times compared to Nb-based ELISA, respectively. This method had LODs of 1.4 μg kg in barley, 0.56 μg kg in oats, and 0.84 μg kg in rice for OTA. The average recovery percent was in a range of 84-137%, and the relative standard derivation percent ranged from 0.64% to 7.8%. The content of OTA in contaminated cereal samples was determined by both the developed Nb-AviTag-based method and liquid chromatography-tandem mass spectrometry (LC-MS/MS). The results demonstrated that the Nb-AviTag was a robust and promising bioreceptor in highly sensitive detection of OTA and other low molecular weight compounds using BA system.

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The aim of the present study was to investigate the protein expression of the autophagy-related genes, BECN1 and PTEN, and the association with drug resistance in epithelial ovarian cancers. In total, 40 patients with pathologically diagnosed epithelial ovarian cancer were divided into a chemotherapy-sensitive group (n=20) and a chemotherapy-resistant group (n=20), according to the results of the pre- or post-operative normative chemotherapy and the post-operative follow-up. The protein expression of the phosphatase and tensin homolog (PTEN) and the BECN1 gene product, Beclin-1, was analyzed using immunohistochemistry in the 40 patients with ovarian carcinoma. The positive rate of Beclin-1 expression was significantly lower in the resistant group (35.0%) compared with the sensitive group (50.0%). The positive rate of PTEN expression was also significantly lower in the resistant group (30.0%) compared with the sensitive group (65.0%). Furthermore, the differences in the expression rates were revealed to be significant (P<0.05). The expression of Beclin-1 was identified to be positively correlated with the expression of PTEN (rs=0.816; P<0.0001). The low expression of the Beclin-1 and PTEN proteins in the ovarian cancer tissues was revealed to be closely associated with drug resistance. Therefore, Beclin-1 may interact with PTEN to participate in the mechanism of drug resistance and the changes in macrophage activity observed in cases of drug-resistant ovarian cancer.