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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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  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The RT-PCR detection of measles virus is more sensitive than the classic virus isolation method, and the result can be obtained within 5-6 hours, but it has the disadvantages of easy contamination and high false positive rate

Method used

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  • Measles virus fluorescent augmentation detection kit and detection method

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Comparison scheme
Effect test

Embodiment 1

[0060] 11 patient gargles (collection time is 3 to 4 days after the rash) were applied to the TaqMan fluorescent quantitative RT-PCR method of the present invention for 2 outbreaks of measles, and the conventional RT-PCR method and virus isolation method were used to detect, wherein the virus 2 were isolated positive, 7 were positive by RT-PCR, and 10 were positive by TaqMan RT-PCR. The positive results of these three tests were consistent. 15 days after the rash appeared, the serum measles virus IgM antibody of the patients was measured. Among the 11 patients, 10 cases of measles IgM antibody were positive, 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 deoxynu...

Embodiment 2

[0075] The measles virus strain Shanghai 191 with known virus titer was serially diluted by 10 times to 1000, 100, 10, 1, 0.1 TCID 50 5 gradients, each gradient takes 200 μL, extracts viral RNA, and detects by fluorescent quantitative PCR (the method is the same as in Example 1). For each concentration of the sample, it is repeated 3 times, and 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 show that the sensitivity of measles virus quantitative RT-PCR is 0.1TCID 50 .

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

[0077]

virus concentration

TCID 50 / Tube

ct value

...

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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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