Nanogold cage kit, preparation method thereof and application of nanogold cage kit in determination of H-FABP
A nano-gold and kit technology, which is applied in the field of biotechnology applications, can solve the problems of low sensitivity and specificity, weak resolution of optical signals, etc., and achieve the effects of high repeatability, improved sensitivity and good stability.
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Problems solved by technology
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Examples
Embodiment 1
[0031] A method for preparing a nanometer gold cage kit, specifically the following steps:
[0032] (1) Preparation of hollow cubic gold nanocages:
[0033] Preparation of HAuCl with deionized water 4 solution at room temperature using a syringe pump in HAuCl 4 Add citric acid dropwise to the solution, and then continue to stir until the color turns red and centrifuge to remove the supernatant to obtain cube hollow nanocrystals of different sizes, then resuspend the cube hollow nanocrystals with deionized water, and centrifuge to remove the supernatant. Much Au 3+ ionized, deionized water to resuspend the precipitate, and add 5mg / ml PVP monomer and 0.1% Tween to obtain a nano gold cage solution; the size of the square hollow nano crystals is 18nm and 40nm.
[0034] (2) Preparation of gold-labeled antibody
[0035] with 0.1M K 2 CO 3 Adjust the pH of the nano-gold cage solution to 8.5; react the specific single-stranded DNA with the nano-gold cage solution for 6-8 hours, ...
Embodiment 2
[0051] (1) Purification of human myocardial H-FABP:
[0052] Human left ventricular myocardium was homogenized in buffer solution, centrifuged in multiple steps and washed with (NH 4 ) 2 SO 4 Carry out salting out→centrifugation→precipitation buffer resuspended and dialyzed protein crude extract→Sephadex G-75 gel filtration→electrophoresis to determine the collection tube number of the main protein bands and collect them→Q Sepharose Fast Flow anion exchange chromatography→polyethylene Diol concentration → buffer dialysis to obtain human heart-type fatty acid binding protein.
[0053] The purified product uses human heart-type fatty acid binding protein from SIGMA Company as a reference standard, and the relative molecular mass is identified by Tricine-SDS-PAGE electrophoresis to 15KD, and the purity is over 90%, which is confirmed by mass spectrometry.
[0054] (2) Preparation of H-FABP monoclonal antibody
[0055] Immune animals: Select 8-12 weeks old Balb / c mice of the s...
Embodiment 3
[0073] Example 3 Urine Test
[0074] 1. Detection object
[0075] Firstly, urine samples from 300 patients with confirmed acute myocardial infarction and acute myocardial injury were selected from the Han population in Nanjing, Jiangsu, and urine samples from 300 healthy subjects were collected as controls to rule out acute myocardial infarction and myocardial injury. .
[0076] 2. Inclusion criteria for case and control samples
[0077] In accordance with the 2007 and 2012 global definition of acute myocardial infarction in China, patients who were hospitalized and diagnosed with acute myocardial infarction underwent cardiac surgery to injure the myocardium.
[0078] 3. Detection method
[0079] Immerse the detection test strip prepared in Example 1 in a fresh urine sample and add it dropwise to the sample hole, observe it in 15 minutes, the standard for interpreting the test result of the test strip is: positive, there are positive marks at the detection line and the qual...
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Abstract
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