Folding primer multiple PCR malaria molecule diagnostic kit and detection method thereof
A folding primer and molecular diagnosis technology, applied in the field of medical testing, can solve the problems of complicated operation, easy contamination and mistakes, long test time, etc., and achieve the effect of enhancing sensitivity and specificity, improving efficiency, and avoiding non-specific amplification.
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Embodiment 1
[0060] Embodiment 1, application folding primer multiplex PCR (FP-PCR) malaria molecular diagnostic kit detects and identifies 4 kinds of human Plasmodium-sensitivity detection, the results are shown in Figure 1-Figure 4 .
[0061] Preparation of blood samples on hemofilter paper simulated by Plasmodium density gradient infection: Plasmodium vivax (PV), Plasmodium falciparum (PF), Plasmodium ovale (PO) (including variants) and Plasmodium malaria ( PM) Infected patients (protozoa density 10000 worms / μl blood or more) contain 1 part of whole blood containing worms, and use EDTA anticoagulated whole blood of healthy people to serially dilute 10 times to contain 10000, 1000, 100, 10 and 1 worm per 1 μl The gradient of worms contains whole blood of worms. Use a pipette to quantitatively drip the Plasmodium density gradient blood filter paper blood sample containing worms (4μl / each 6mm diameter filter paper disc), and after natural air drying, divide into sealed tubes and store at...
Embodiment 2
[0078] Embodiment 2, application folding primer multiplex PCR (FP-PCR) malaria molecular diagnostic kit detects and identifies and simulates 4 kinds of human Plasmodium mixed infections, the results are shown in Figure 5 .
[0079] Preparation of blood samples on blood filter paper for simulating mixed infection of 4 kinds of Plasmodium: the possible density of different species of plasmodium mixed infection may vary greatly. Considering the importance of detecting low-density PF among them, use the density gradient filter paper of 4 kinds of parasites prepared in Example 1 The blood sample template simulates the mixed infection of 3 groups (respectively 2, 3 and 4 kinds of protozoa), and each group has 4 combinations, and a total of 12 mixed templates with different combinations of insect species and density are prepared. The highest and lowest values of each combination Densities vary by a factor of 100. Half of the combinations use PF as the lowest density, and the othe...
Embodiment 3
[0086] Embodiment 3, specificity determination
[0087] According to the operation method of Example 1, 10 healthy human blood samples were tested repeatedly 7 times, and there was no amplified band except dimer below moderate. Therefore, it will not affect the detection results of malaria infection.
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