Application of Tris |CAS:77-86-1| in COVID-19 Detection Kit

Tris is an important biological buffer with a pKa of 8.06 at 25°C and a pH buffer range of 7.0-9.0. It is widely used in the preparation of buffers in the fields of biochemistry and molecular biology. In the CN111551743A patent, the application of Tris in the COVID-19 detection kit is introduced. The kit consists of 6 parts, specifically: luminescence substrate solution A, luminescence substrate solution B, sample dilution solution, enzyme labeling working solution, calibrator and magnetic bead suspension. I will briefly describe the composition and preparation method of each component .

(1) Luminescent substrate solution A

The luminescent substrate solution A consists of three parts: Tris, hydrochloric acid and luminol;

Preparation method: Weigh the components, dissolve them in distilled water respectively, and set the volume to constant volume; wherein Tris 0.01-0.05g/L, hydrochloric acid 0.002-0.01ml/L, luminol 0.001-0.007g/L;

(2) Luminescent substrate solution B

The luminescent substrate solution B is composed of four parts: sodium dihydrogen phosphate, disodium hydrogen phosphate, carbamide peroxide and sodium chloride;

Preparation method: Weigh each component, dissolve it in distilled water respectively, and set the volume to obtain; wherein sodium dihydrogen phosphate 0.1-0.5g/L, disodium hydrogen phosphate 2.0-4.0g/L, carbamide peroxide 0.002-0.05 g/L, sodium chloride 3-15g/L.

(3) Sample Diluent

The sample diluent consists of three parts: BSA (bovine serum albumin), sodium chloride and ProCLin300.

Preparation method: Weigh each component, dissolve it in distilled water, make up to volume, and get it; wherein BSA 8-15g/L, sodium chloride 5-12g/L, ProCLin300 0.1-1ml/L;

(4) ELISA working solution

The enzyme-labeled working solution is consists of six parts：nCov-Ag-HRP, sodium dihydrogen phosphate, disodium hydrogen phosphate, aminopyrine, ProCLin300 and CaSein.

Preparation method: Weigh each component, dissolve it in distilled water, and make up to volume. Among them, nCov-NAg-HRP 0.3-1.0mg/L, nCov-SAg-HRP 0.8-1.5mg/L, sodium dihydrogen phosphate 0.05-0.4g/L, disodium hydrogen phosphate 2.0-4.0g/L, aminopyrine 0.2-1.5g/L, ProCLin300 0.1-1.0ml/L, CaSein 1.0-4.0g/L;

(5) Calibrator

The calibrator used in the patent is human serum.

(6) Magnetic bead suspension

Magnetic bead suspensions include magnetic microparticles conjugated to anti-human IgM antibodies.

Preparation method: Weigh magnetic particles, activate with carbodiimide (EDC), add anti-human IgM antibody, suspend, wash, and make up to volume. Magnetic particles coupled with anti-human IgM antibody (in terms of the mass of the magnetic particles) 0.2-2g/L, wherein 1g of the magnetic particles is coupled with 1-2g of anti-human IgM antibody;

The kit prepared by the above method has the following beneficial effects:

① The kit uses the N protein and S protein fragments of COVID-19 as antigens together, and the combined use of the two improves the detection rate, sensitivity and specificity of the kit.

②The kit specifically selects the type of anti-human IgM antibody conjugated on the magnetic particles, and it is found that the mouse anti-human IgM antibody has a relatively better effect and a higher positive detection rate when used in the present invention.

References

CN111551743A, Beijing Bell Bioengineering Co., Ltd.