Measles virus fluorescent augmentation detection kit and detection method

A technology for detecting kits and measles virus, applied in biochemical equipment and methods, microbe measurement/inspection, etc., can solve the problems of easy pollution and high false positive rate in detection, and achieve convenient and accurate judgment, high sensitivity, and quantitative analysis Effect

Inactive Publication Date: 2007-01-31
ZHEJIANG CENT FOR DISEASE CONTROL & PREVENTION
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Problems solved by technology

The RT-PCR detection of measles virus is more sensitive than the classic virus isolation method, and the result

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  • Measles virus fluorescent augmentation detection kit and detection method

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Example Embodiment

[0059] Example 1:

[0060] 11 patient gargles from 2 measles outbreaks (collection time is 3 to 4 days after rash) were detected using the TaqMan fluorescent quantitative RT-PCR method, conventional RT-PCR method and virus isolation method of the present invention. Two copies were positive for separation, 7 copies were positive for RT-PCR, and 10 copies were positive for TaqMan RT-PCR. The results of the positive patients tested by these three were the same. After 15 days of eruption, the serum measles virus IgM antibody was measured, and 10 of 11 patients showed positive measles IgM antibody, which was completely consistent with the results of TaqMan RT-PCR.

[0061] The TaqMan RT-PCR reaction system is as follows:

[0062] 2×RT-PCR buffer 12.5μL

[0063] RNase inhibitor RNasin 0.8 activity unit / μL

[0064] Specific amplification upstream primer 0.4μM

[0065] Specific amplification downstream primer 0.4μM

[0066] Specific probe 0.2μM

[0067] The mixture of deoxynucleoside trip...

Example Embodiment

[0074] Example 2:

[0075] The measles virus strain Shanghai 191 with known virus titer is diluted by 10-fold series to 1000, 100, 10, 1, 0.1 TCID 50 5 gradients, 200μL for each gradient, extract viral RNA, perform fluorescence quantitative PCR for detection (the method is the same as in Example 1), repeat 3 times for each concentration of the sample, the results are shown in figure 1 . The composition of the fluorescent quantitative PCR reaction system is the same as in Example 1. The results showed that the sensitivity of measles virus fluorescence quantitative RT-PCR was 0.1TCID 50 .

[0076] The repeatability of the method is evaluated by evaluating the coefficient of variation (CV) of the Ct value of each dilution repeated three times. The results are shown in Table 1. The coefficient of variation is less than 3%, indicating that the method has good repeatability.

[0077]

[0078] The measles virus strain Shanghai 191 is diluted by 10-fold series to 1000, 100, 10...

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Abstract

The present invention provides a kind of measles virus fluorescence amplification detection kit and method for detecting measles virus by using said kit. Said kit includes measles virus hemagglutinin gene standard product, fluorescence amplification detection reagent and DNA polymerase and reverse transcriplase. The fluorescence amplification detection reagent mainly includes one-step method RT-PCR buffer fluid, specific amplification primer and specific probe and deoxynucleoside triphosphate mixture. Said method can directly detect pathogen nucleic acid, and can detect the specific gene containing measles virus.

Description

(1) Technical field [0001] The invention relates to a rapid gene detection kit and detection method of measles virus. (2) Background technology [0002] Measles is an acute infectious disease caused by measles virus, characterized by fever, respiratory symptoms and rash. Since the Pan American Health Organization (PAHO) first put forward the strategic goal of measles elimination in the World Health Organization (WHO) Americas Region in 2000, the global measles elimination activities have made substantial progress. In order to speed up the control of measles, the General Office of the Ministry of Health issued the "National Measles Surveillance Program (Trial)" in 1998, requiring that in addition to measuring serum measles IgM antibodies, it is also proposed to isolate measles virus or reverse transcription-polymerase chain reaction (RT-PCR) RT-PCR) method to confirm the diagnosis of measles cases. The RT-PCR detection of measles virus is more sensitive than the classic vir...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 卢亦愚严菊英冯燕茅海燕徐昌平龚黎明
Owner ZHEJIANG CENT FOR DISEASE CONTROL & PREVENTION
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