Novel coronavirus antigen and application thereof
An antigen and antibody technology, applied in the direction of virus, virus peptide, virus/bacteriophage, etc., can solve the problem of poor specificity in the detection of new coronavirus, achieve the effect of improving specificity and avoiding false positives
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Embodiment 1
[0023] 1.1 Using bioinformatics methods, combined with previous experience, screen out the following preferred S segment of SARS-CoV-2:
[0024] 123-ATNVVIKVCEFQFCNDPFLGVYYH-146 (SEQ ID NO.1)
[0025] 209-PINLVRDLPQGFSALEPLVDLPIGI-233 (SEQ ID NO.2)
[0026] 262-AAAYYVGYLQPRTFLLK-278 (SEQ ID NO.3)
[0027] 485-GFNCYFPLQSYGF-497 (SEQ ID NO.4)
[0028] 504-GYQPYRVVVLSFELLHAPATVCGPK-528 (SEQ ID NO.5)
[0029] 1.2 The preferred N fragment of SARS-CoV-2 obtained by screening is as follows:
[0030] 52-WFTALTQHGK EDLKFPRGQG VPINTNSSPD DQIGYYRR-89 (SEQ ID NO. 6)
[0031] 106-PRWYFYYLGT GPEAGLPYGA NKDGIIWVAT EGALNTPKDH (SEQ ID NO. 7)
[0032] IGTRNPANNA AIVLQLPQGTT-166 (SEQ ID NO. 8)
[0033] 217-AALALLLLDRL-227 (SEQ ID NO.9)
[0034] 243-GQTVTKK-249 (SEQ ID NO.10)
[0035] 267-AYNVTQA-273 (SEQ ID NO. 11)
[0036] 1.3 Connect the above polypeptides to each part through the GGGGS connecting peptide, and the specific amino acid sequence is as follows:
[0037] ATNVVIKVCE FQFCND...
Embodiment 2
[0068] Example 2 The N1 protein prepared in Example 1 was used to immunize animals
[0069] 2.1 Immunize rabbits with purified N1 protein respectively, take about 400 μg of purified N1 protein and mix completely with an equal volume of Freund's complete adjuvant, and immunize long-eared white rabbits. Subcutaneous injection was used for immunization. After 4 weeks, the same dose of Freund's incomplete adjuvant was used to boost immunization once. After the booster immunization, blood was collected once every 2 weeks, 4 weeks, and 6 weeks respectively to obtain rabbit polyantiserum R1.
[0070] 2.2 Immunize rabbits with 200 μg each of purified N2 and S2 proteins, mix completely with an equal volume of complete Freund's adjuvant, and immunize long-eared white rabbits. Subcutaneous injection was used for immunization. After 4 weeks, the same dose of Freund's incomplete adjuvant was used to boost immunization once. After the booster immunization, blood was collected once every 2 w...
Embodiment 3
[0073] Embodiment 3 prepares kit
[0074] 3.1 The diluted N1 protein was used as the coating antigen, and the HRP-labeled N1 protein was used as the labeled antigen to detect the total antibody, and the kit was named K1.
[0075] 3.2 According to the method in 3.1 above, using N2-N8 proteins as coating antigens and HRP-labeled N2-N8 proteins as labeled antigens, prepare corresponding kits K2-K8 for detecting N2-N8 proteins respectively.
[0076] 3.3 According to the method in 3.1 above, using S2-S8 proteins as coating antigens and HRP-labeled S2-S8 proteins as labeled antigens, prepare corresponding kits K9-K15 for detecting S2-S8 proteins respectively.
[0077] 3.4 The detection steps of the above kits are as follows (taking kit K1 as an example): Prepare coated plates with N1 purified protein, and use sample diluent (containing BSA, Tris-Cl and Tween-20) for serum samples at a ratio of 1:100 Dilute, add 100 μL to each well, incubate at 37°C for 0.5 h, wash each well with PB...
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