Abstract: Objective To develop a blood-safety test method based on detecting nucleic acids of a pathogen and visually to determine which kind of pathogens (HBV, HCV, HIV, or TP) in the blood. Methods The nucleic acids of four pathogens were amplified with long-primer PCR. The steps of PCR cycles were simplified from traditional three steps to two steps. PCR products labeled with antigens were applied to the gold-nanoparticles dipstick immobilized with antibodies. The developing results were determined by naked eyes. The annealing temperatures and denaturing temperatures for the four pathogens were optimized and the accuracy of the method for detecting the stimulated and the serum samples of four pathogens were evaluated.Results The annealing temperatures of four pathogens were 76°C, 78°C, 76°C and 78°C; the denaturing temperatures of them were 84°C, 90°C, 84°C and 88°C. The assay was successfully developed for discriminating the positive and negative samples with high accuracy. The amplification time was only 40 min/40 cycles and the detection time was 10 min. Conclusion The assay which developed in this study can be used for detecting nucleic acids of four pathogens visually. In comparison with traditional methods, the assay has the advantages of the rapidity, low cost and no requirements of special instruments and holds a great promise for wide applications in small hospitals.
黄欢，李朔，孙丽洲，周国华
. Development of a Long-Primer Fast PCR Coupled with Visual Nucleic Acid Dipstick Assay for Detection of Four Pathogens[J]. Labeled Immunoassays and Clinical Medicine, 2013
, 20(6)
: 436
-440
.
DOI: 10.11748/bjmy.issn.1006-1703.2013.06.022
