| Hepatitis B, a liver disease associated with hepatitis B virus (HBV), is a big health problem for human beings. It is is widely distributed over the world and millions of people are suffered from it. In China,300,000 people die from chronic liver disease each year and about 130,000 of them are the victim of primary liver cancer. What makes matters worse is that HBV could be transmitted to infants by their mother carrying the virus in a possibility of 30%. Infants are apt to be infected with HBV and become hepatitis B carriers because of their imperfect immunity. After growing up, they often gradually become victims of chronic hepatitis, cirrhosis, and primary hepatocellular carcinoma. In order to prevent the transmission of HBV, it’s essential to develop a rapid and accurate diagnosis to detect the infection of HBV.Firstly, a rapid diagnostic kit to detect HBsAg with a cutoff value of 2.0 ng/ml was developed based on immune colloidal gold technique and double antibody sandwich technique. Concentrations of the primary antibody coated with colloidal gold and the second antibody in the test line (T-line) were investigated and optimized. Optimization of nitrocellulose membrane types, volume of the second antibody in the T-line, and composition of the sample pad solution were conducted. The optimal values for corresponding parameters were as following; the coating pH was 7.0; the coating ratio of colloidal gold solution and antibody was 100 ml/mg; the type of nitrocellulose membrane was Sartorius; the concentration and dosage of the second antibody concentration was 1.0-1.5 mg/ml and 2.0 μl/cm, respectively; the sample pad solution was 0.1 M of Tris-HCl solution containing 1% of Triton X-100,0.5% of PVP, and 0.1% of BSA with pH of 10.0.Secondly, performance verification analysis, risk analysis and process verification analysis were perfomed to verify the overall performance of the developed kit. The sensitivity, specificity, detection range, accuracy, and within-run imprecision inaccuracy were examined to evaluate the performance of the kit, and risk analysis and process verification analysis were combined to achieve an overall evaluation of the product quality. Results exhibited that all the performance indexes of the products had meet the requirements. |
