Anti-human respiratory syncytial virus N protein antibodies and immunochromatographic kit using the same
A technology of syncytial virus and immune chromatography, applied in the field of biomedicine, can solve the problems of low specificity, low titer, low purity, etc.
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Embodiment 1
[0071] Embodiment 1 Preparation of two kinds of rabbit anti-human respiratory syncytial virus N protein antibodies
[0072] The preparation methods of two kinds of rabbit anti-human respiratory syncytial virus N protein antibodies are as follows:
[0073]1) After structural biological analysis and related experimental research, a short peptide composed of 14 amino acids at positions 21-34 and 14 amino acids at positions 226-239 of human respiratory syncytial virus N protein (GenBank sequence number AAB59852.1) was selected As two linear epitopes for preparing rabbit anti-human respiratory syncytial virus N protein antibody respectively, the two amino acid sequences are SKYTIQRSTGDSID and FGIAQSSTRGGSRV respectively, and these two sequences are respectively named N1 and N2;
[0074] 2) After adding a cysteine to the C-terminal of N1 and the N-terminal of N2 of the amino acid sequence described in step 1), respectively synthesize and purify the polypeptides with an automatic p...
Embodiment 2
[0081] Example 2 Preparation and Application of Immunochromatography Kit Based on Quantum Dot Labeling Technology
[0082] 1. Quantum dot-labeled antibody AbN1
[0083] Add 0.4nmol carboxyl water-soluble quantum dots and 800nmol carbodiimide (EDC) to the microcentrifuge tube successively, make the volume to 1ml with MES buffer (10.66g / LMES, 0.74g / LEDTApH7.4), and keep mixing solution, reacted at 37°C for 5 minutes, then added 0.34 mg of the antibody AbN1 prepared in Example 1, and reacted in the dark for 2 hours, and added single-terminal amino polyethylene glycol (PEG2000-NH2) to a final concentration of 1% (m / v ), block the unreacted activated carboxyl site, and continue to react in the dark for 1 h. The reacted sample was centrifuged with an ultrafiltration tube (molecular weight cut-off 100k) at 6500rpm for 5min to a volume of 200ul. The ultrafiltered sample was transferred to an ordinary EP tube and centrifuged to remove aggregates (10000rpm, 3min). Add the supernatant ...
Embodiment 3
[0106] Example 3 Preparation and Application of Immunochromatography Kit Based on Colloidal Gold Labeling Technology
[0107] 1. Colloidal gold-labeled antibody AbN1
[0108] a. Preparation of 30nm colloidal gold
[0109] Take a siliconized 250ml Erlenmeyer flask, add 99ml ultrapure water, add 1% (m / v) HAuCl4 solution therein and mix evenly, heat in an oil bath and stir until boiling. 2ml of 1% (m / v) trisodium citrate aqueous solution was quickly added thereto, and the solution continued to boil for 10 minutes (the solution turned from blue to red during this process). Stop heating, let the solution cool down to room temperature naturally, then add ultrapure water to it to make up to 100ml.
[0110] b. Colloidal gold-labeled antibody AbN1
[0111] 1) Take a siliconized 50ml triangular flask, add 10ml of the colloidal gold solution prepared in step a, add 240ul0.2mol / LK to the gold solution 2 CO 3 Adjust the pH to 8.5;
[0112] 2) Add the antibody AbN1 into the colloidal ...
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