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Information on the preparation of colloidal gold solutions for the Antigen test

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Information on the preparation of colloidal gold solutions for the Antigen test

The preparation of a colloidal gold solution for the Antigen test adopts the trisodium citrate reduction method to prepare a colloidal gold solution. Take 1000 mL of ultrapure water and heat it to boiling on a magnetic stirrer, add 40 mL of 1% chloroauric acid solution, when the solution boils again, quickly add 36 mL of 1% trisodium citrate solution, continue to boil when the color of the solution turns purple 5min, turn off the heater, and restore to the original volume with ultrapure water after cooling. The average diameter and Zeta potential of colloidal gold particles were measured with a nanoparticle size analyzer and stored at room temperature in the dark. So what is the information about the preparation of colloidal gold solutions for the Antigen test? Let's take a look.

 

Here is the content list:

l Preparation of Colloidal Gold Labeled Conjugates

l Preparation of rapid Antigen test strips

l Minimum detection limit

l Stability test

 

Preparation of Colloidal Gold Labeled Conjugates

Take 100 mL of Antigen test colloidal gold into two clean beakers, respectively, according to the optimized marked pH conditions, add 3.3 mL and 2.1 mL of 0.1 mol/L potassium carbonate to adjust the pH, stir with a magnetic stirrer at 300 r/min for 10 min, and add Mouse anti-N protein monoclonal antibody and DNP-BSA protein were each 1 mg and stirred for 15 minutes; 2 mL of 10% BSA solution was added, and 2 mL of 10% BSA solution was added, and the stirring was continued for 20 minutes; after stirring, the Antigen test colloidal gold solution was transferred to two 50 mL centrifuge tubes, set the centrifuge parameters were 9000 r/min, 20 min, and 4 °C, and centrifugation was performed. After centrifugation, take out the centrifuge tube carefully, do not shake, remove the supernatant with a pipette, collect the precipitate, reconstitute it with conjugate diluent to 10 mL, and store at 4°C in the dark. The average diameter and Zeta potential of the Antigen test colloidal gold conjugate particles were measured by a nanoparticle size analyzer.

 

Preparation of rapid Antigen test strips

The sample pad was uniformly coated with the sample pad treatment solution, and the coating amount of each glass fiber was 30 mL and dried at 50°C for 24 hours. Dilute the rapid Antigen test colloidal gold-labeled mouse anti-N protein monoclonal antibody and colloidal gold-labeled DNP-BSA protein conjugate with conjugate diluent, the volume ratio is 18% and 20%, respectively, and treat with the diluted colloidal gold conjugate the coupler pad was uniformly coated with liquid, the coating amount of each glass fiber was 30mL, and it was dried at 45°C for 24h. The mouse anti-N protein monoclonal antibody and the rabbit anti-DNP polyclonal antibody were diluted with PBS containing 0.01mol/L pH7.4 and 5% trehalose to the concentration of 2.0mg/mL and 1.0mg/mL, respectively. As the detection line (T line) and the quality control line (Cline), the capture antibody was coated on a nitrocellulose membrane (NC membrane) and dried at 45°C for 48h. The prepared NC film, conjugate pad, sample pad, and water-absorbing pad were pasted on the PVC bottom plate in turn, cut into 3mm/stripe with a slitter, put into a card case, and packaged in an aluminum foil bag. During the test, insert the nasopharyngeal swab into the lifting tube pre-filled with the extraction solution (10mmol/LPBS, pH7.4, 1%NP-40), and add 3 drops (100μL) dropwise to the sample hole of the test card after extraction for 15s. 15min reaction time for visual interpretation. Antigen test T line and Cline are both positive, T line is not colored, Cline is negative, Cline is not colored, indicating that the test strip is effective, and it needs to be re-tested.

 

Minimum detection limit

Minimum detection limit studies were performed with heat-killed virus cultures. The eluate from nasopharyngeal swabs from healthy people was used as the negative matrix for the Antigen test. First, a 10-fold gradient dilution was performed. After the negative result of the rapid Antigen test, the previous concentration was then 2-fold gradient dilution, and no less than 19 times out of 20 tests were positive. The concentration level (≥95%) was used as the minimum detection limit of the reagent.

 

Stability test

Place the prepared Antigen test finished reagent card in a 50°C oven, and test with high and low concentration quality control substances every two weeks. Repeat the test 3 times for each sample. The difference in the band depth between the high temperature accelerated destruction reagent and the control reagent for 1 week and 8 weeks.

 

The above is the relevant information about the preparation of the colloidal gold solution for the Antigen test. If you are interested in the COVID-19 rapid Antigen test, SARS-CoV-2 Rapid Antigen test, you can contact us. Our website is www.bioteke.cn.


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