Preliminary comparison of six commercially available fluorescence PCR kits in detecting nucleic acid of enterovirus
Objective To compare preliminarily the sensitivity and specificity of EV-A71/CVA16/enterovirus(EV)nucleic acid detection kits and CVA6/CVA10 nucleic acid detection kits of six commercial brands,so as to provide reference for selection of detection reagents as well as disease prevention and surveillance.Methods The virus strains of four serotypes,including EV-A71,CVA16,CVA6 and CVA10 were selected as the sample pool.The sample strains were serially diluted in ten fold.The nucleic acids of serial diluted samples were quantified by digital PCR.The samples were then mixed to created a pool of mixture and diluted in serial.The real-time fluorescence PCR was then used for detection.The detection limit and sensitivity were analyzed.The kits were used to analyze nucleic acid from virus other than enterovirus to evaluate the specificity.Group scoring and quantity scoring were used to evaluate the characters of amplification curves and Ct values and obtain the generalized scores.Results In the comparison of sensitivities,kit A,D and E were more sensitive.In the analysis of amplification curve and Ct values,the top three kits in universal detection of enteroviruses were kits A,C and E and kits for detection of EV-A-71,CVA16,CVA6 and CVA10 were kits E,A and F,kits A,D and E,kits E,D and F,and kits E,A and C,respectively.All kits had good specificity.Conclusions In the triplex fluorescence PCR kits for detecting nucleic acid of EV-A71/CVA16/EVs,the kits A,E,C and D had higher general scores.For duplex fluorescence PCR kits for detecting nucleic acid of CVA6/CVA10,kits E and F had higher general scores.It is recommended to have kits of two or more brands and compare the detection results.The quantitative scoring method could be the reference for the evaluation of other fluorescence PCR kits.
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