Effect of different diluents on stability of mixed enzyme-labeled antibody / 中国生物制品学杂志

@#Objective To investigate the effect of different diluents on the stability of the mixed enzyme-labeled antibody,and screen a suitable diluent for the mixed enzyme-labeled antibody,which can protect the stability of the cocktail mixture of horseradish peroxidase(HRP) conjugated secondary antibody and alkaline phosphatase(AP) conjugated secondary antibody.Methods Using Tris-HCl buffer as the base solution,four different diluent formulations(A,B,C,D) were prepared with different kinds of stabilizers contained in each formula,such as protein,metalion,surfactant and bacteriostatic agent.Mixed enzyme-labeled antibodies were prepared with different diluents and stored at 2～8 ℃ and 37 ℃,which were detected for the stability by ELISA,enzyme activity assay and immunohistochemistry(IHC) staining.Results D solution [Tris(50 mmol/L)+BSA(1.5%,g/100 mL)+Tween-20(0.3%)+Proclin-950(0.1%,g/100 mL)+sodium chloride(150 mmol/L)+L-Ascorbic Acid(1 mmol/L)+L-methionine(5 mmol/L)+L-histidine(5 mmol/L)+sodium caseinate(1%,g/100 mL)+zinc chloride(0.1 mmol/L)+trehalose(8%,g/100 mL)] was of the optimal protective effect.When stored at 37 ℃ for 10 weeks,the working solution of HRP conjugated secondary antibodies and AP conjugated secondary antibodies showed the highest titer,the activity residual ratios of HRP and AP were 93.46% and96.07%,respectively,and there was no significant difference in IHC results.Conclusion This formula diluent can be used for the preparation of mixed enzyme-labeled antibody working solution.

Comparison of an Artificial Neural Network and a Response Surface Model during the Extraction of Selenium-Containing Protein from Selenium-Enriched Brassica napus L.

In this study, the extraction conditions for selenium-enriched rape protein (SEP) were optimized by applying a response surface methodology (RSM) and artificial neural network (ANN) model, and then, the optimal conditions were obtained using a genetic algorithm (GA). Then, the antioxidant power of the SEP was examined by using the DPPH, ABTS, and CCK-8 (Cell Counting Kit-8), and its anticancer activities were explored by conducting a cell migration test. The results showed that compared with the RSM model, the ANN model was more accurate with a higher determination coefficient and fewer errors when it was applied to optimize the extraction method. The data obtained for SEP using a GA were as follows: the extraction temperature was 59.4 °C, the extraction time was 3.0 h, the alkaline concentration was 0.24 mol/L, the liquid-to-material ratio was 65.2 mL/g, and the predicted content of protein was 58.04 mg/g. The protein was extracted under the conditions obtained by the GA; the real content of protein was 57.69 mg/g, and the protein yield was 61.71%. Finally, as the concentration of the selenium-containing protein increased, it showed increased ability in scavenging free radicals and was influential in inhibiting the proliferation and migration of HepG2 cells.