6. Protein Silver Stain Kit, 25T (Protein Silver Staining Kit) $ 448

Product Information

Description

Silver staining is a highly sensitive staining method, the basic principle of which is to display the protein band by forming a black precipitate on the protein by the reduced anion. The silver stain is 100 times more sensitive than the traditional Coomassie brilliant blue stain and can detect proteins of less than 0.5 ng. Because of its high sensitivity, silver staining is widely used in 2D gel analysis and vertical PAGE gel for very low protein content determination. It is a common method for the detection of low abundance proteins in gels. This product can be used for protein detection in SDS-PAGE or PAGE polyacrylamide gel, with high sensitivity, easy to operate, almost colorless background, silver staining of one gel can be completed within 90 min. This kit can stain 25 gels of the same specification.

Storage and Handling Conditions

Transport at room temperature; The sensitizer was stored at room temperature, and the other reagents were stored at 4℃, and the validity period was 12 months.

Component

Assay Protocol

1. Fixation: Prepare fixation solution according to the table below. Put the gel in a clean dye VAT, clean it with ultra-pure water for three times, add 20 mL of fixing solution to cover the gel, and slowly shake it with a horizontal shaker (60-70 rpm) and incubate it for 30 min. In order to prevent acetic acid and ethanol volatilization, please seal the dye VAT with plastic wrap. Prolonged fixation time can further reduce the background.

2. Washing: Pour out the fixing solution, replace the ultra-pure water covering gel, and shake the horizontal shaker for three times, 10 min each time.

3. Sensitization: Prepare the sensitization working solution according to the table below. The sensitization solution was added to cover the gel, and the gel was incubated for 1 min with slow shaking on a horizontal shaker.

4. Washing: Pour out the sensitization working solution, replace with ultra-pure water, and shake the shaker for three times, 20 s each time.

5. Silver dyeing: Prepare silver dyeing solution according to the following table. Silver staining solution was added to cover the gel, and the gel was incubated for 20 min by shaking slowly. Silver dyeing solution should be prepared and used now, and should be used within 2 hours as far as possible.

6. Wash with water: discard the dye solution, replace with ultrapure water, and shake the shaker for three times, 20 s each time.

7. Color development: Prepare the developing solution according to the table below. The developing solution was added to cover the gel, and the gel was incubated with shaking for 1-5 min until the ideal target protein band appeared.

8. Termination: Prepare the development termination solution according to the table below. The development stop solution was added to cover the gel, and the gel was incubated for 2 min with a shaker. It is normal for gas to be produced during the process, and the gas produced is CO2. Then the stop solution was discarded and the gel was washed with ultrapure water for 2-5 min.

9. Storage: After cleaning, the gel can be placed for observation and photography. The stained gels can be stored in ultrapure water or 1% acetic acid solution, or dried in an appropriate manner.

Note:

1. To prevent interference from impurities, use high purity water (resistance > 16 MΩ/cm) and clean glassware or plastic containers. Do not use metal containers.

2. Please take protective measures during the whole experiment. Ultrapure water, absolute ethanol and glacial acetic acid should be prepared by yourself.

3. Do not directly touch or press the colloid during the operation, and the dye VAT used for dyeing cannot use metal products, glass or plastic containers are the best.

4. Keep the sensitizer at room temperature away from light. Do not store it at 4℃, otherwise it will fail.

5. All working fluids shall be prepared and used at present. After opening and using the reagent, it is necessary to tighten the cap of the bottle to prevent the volatilization of the solution and chemical reaction with substances in the air, which will affect the dyeing effect.

6. The developing solution is stored at 4℃ for a long time with crystal precipitation (normal phenomenon), and can be incubated at 37℃ until completely dissolved before use.

7. When the gel is thick or the operating temperature is low, the color development time can be extended appropriately.

8. The dyeing time, washing time, color development and termination time of each step should be well controlled, otherwise the glue background will become dark and it is not easy to observe the target band.

For Research Use Only!