Development and verification of double antibody sandwich ELISA for determination of ovalbumin / 中国生物制品学杂志

@#Objective To develop and verify a double antibody sandwich ELISA detection method for the determination of ovalbumin(OVA),in order to determine the OVA content in influenza vaccines.Methods The rabbit anti-OVA polyclonal antibody was used as coating antibody and HRP labeled rabbit anti-OVA polyclonal antibody as detection antibody to develop a double antibody sandwich ELISA for OVA.The antibody concentration(2,1,0.5 and 0.25 pg/mL) for coating,enzyme-labeled antibody concentration(0.5,0.25 and 0.125 μg/mL),and kinds of blocking reagent(blocking with1% BSA,blocking with 2% BSA,blocking with 1% BSA and 1% sucrose,and blocking with 2% BSA and 2% sucrose,using nonblocking as control) were optimized,and the Cut-off value was determined as the judgment standard.The developed method was verified for the linear range and detection limit,specificity,repeatability and accuracy.Results The optimum detection conditions were as follows:the concentration of coating antibody was 1 μg/mL,the concentration of enzyme-labeled antibody was 0.25 μg/mL;The blocking reagent was 2% BSA and 2% sucrose;The Cut-off value was 0.051 66.The linear range of the method was 5～0.313 ng/mL,and the detection limit was 0.078 ng/mL;It did not react with influenza virus,bovine serum albumin(BSA) and Vero cell supernatant;The intraplate and interplate coefficient of variation(CV) were between 2.562%～13.887% and 4.000%～16.497% respectively;The coincidence rate between the results of this method and the Germany SERAMUN OVA quantitative test kit ranged from 93.79% to 107.05%.Conclusion The developed OVA double antibody sandwich ELISA has good specificity,repeatability,linear range and accuracy with convenience and lower cost,which might be used for the quantitative detection of OVA.

Flavonoids Extracted from Fruit Pulp of Cucumis metuliferus Have Antiviral Properties.

Aim: To investigate the antiviral property of flavonoids from Cucumis metuliferus fruit pulp in chicken embryo fibroblast (CEF) cells and embryonated chicken eggs (ECE) induced with infectious bursal disease virus (IBDV). Study Design: Extraction and administration of bioactive extract. Place and Duration of Study: Department of Pharmacology, University of Jos, Nigeria and Virology Department, National Veterinary Research Institute, Vom, Nigeria between June, 2011 and August, 2011. Methodology: The CEF cells were first exposed to 100, 50, 25, 12.5, 6.25, 3.125, 1.563, 0.782, 0.391 and 0.195 mg/ml of the sterile flavonoids to test for cytotoxicity and the cells monitored visually daily using a light microscope for evidence of cytopathic effects at 24, 48 and 72 hours. Toxicity of flavonoids in embryonated eggs and antiviral assay for flavonoids using IBDV were then carried out. Hemagglutination test for antigenicity of the virus was also performed to confirm antiviral activity. Results: The flavonoids (100 to 0.195 mg/ml concentrations) were not cytopathic when exposed to CEF cells. After 24 and 48 hours, all the embryonated eggs died at 100 and 50 mg/ml of the flavonoids respectively, but mortalities were not recorded at other concentrations of the flavonoids. Concentrations of the flavonoids at 100, 50, 25, 12.5 and 6.25 mg/ml were found to be toxic against IBDV, but viral replication was not inhibited from flavonoids concentrations of 3.125, 1.563, 0.782, 0.391 and 0.195 mg/ml. Conclusion: This investigation revealed that flavonoids from Cucumis metuliferus fruit pulp were relatively safe in chickens and possess antiviral activity against IBDV.

Study of production of inactivated influenza vaccine for human on egg-grown from reassortants NIBRG-14 at vaccine institute

Background: A/H5N1 influenza virus spreads from birds to humans and cause influenza diseases with high mortality rate. Vaccination is the most effective way to protect communities from pandemic, reduce morbidity and mortality. The study of creating A/H5N1 influenza vaccines in conformity with Vietnam was the urgent need. Institute of Vaccine\u2019s Achievement (IVAC) studied production of inactivated influenza vaccine for human on egg-grown from reassortants NIBRG-14. Objectives: In order to produce experimentally A/H5N1 influenza vaccine for human in accordance with WHO requirements and set up a viable process for production of the vaccines. Subjects and method: 10 days embryonated eggs and NIBRG-14 strains were served to the study with LAL method to check endotoxin, Kijehdal method to test total protein. Results: IVAC had produced successfully 5 lots of absorbed vaccine A/H5N1 (FLUVAC) using NIBRG-14 strains and embryonated eggs. Initially, production and quality control processes had been set up at IVAC by applying the recommendations of WHO. Conclusion: The success of the study was a basis of the approval of the government to establish a influenza vaccine manufacturing facilities.