Coomassie Brilliant Blue G250, 250 mL (for Protein Quantification)

Product Information

Product Description/Introduction

Coomassie Brilliant Blue G250 can bind proteins rapidly. The maximum absorption of Coomassie Brilliant Blue G250 is 488 nm in the free state. After binding to protein, the maximum absorption changed to 595 nm. The optical absorption value is proportional to the protein content, so it can be used for the quantitative detection of protein. This product can also be used as a supplement to the G2001 Bradford Assay protein quantitative detection kit.

Storage and Shipping Conditions

Ship and store at room temperature, valid for 12 months.

Assay Protocol/Procedures

1. (Optional) Drawing standard curve (microplate reader method): Dissolve BSA in water to prepare 0.5 mg/mL protein standard working solution. The protein standard working solution is added to the 96-well plate at levels of 0, 1, 2, 4, 8, 12, 16, 20 μL, and then the gradient working solution is supplemented to 20 μL with PBS or saline. The gradient curves of protein concentration are 0, 25, 50, 100, 200, 300, 400 and 500 μg/mL. If only relative quantitative comparisons are made, no standard curves are required.

2. Prepare the sample to be tested: The protein sample to be tested is diluted appropriately (the protein concentration of the sample can be detected by pre-experiment to be within the range of the standard curve, to ensure the reliability of the test result), and 20 μL of each sample is added to the 96-well plate. The sample to be tested should be diluted in the same solution as the protein standard.

3. Detection: 200 μL Coomassie brilliant blue G250 solution is added to each well and thoroughly mixed (the 96-well plate can be placed on the oscillator for 30 s). After 3-5 min at room temperature, the standard curve No. 0 is used as a reference, and the colorimetric measurement is performed at 595 nm wavelength, and the absorbance value of each well is recorded.

4. Calculation: The gradient protein content (μg/mL) in the standard curve is taken as the abscissa, and the light absorption value is taken as the ordinate to draw the standard curve. According to the absorbance value of the sample, the protein concentration of the sample to be measured in the corresponding well can be found on the standard curve (μg/mL), and then multiplied by the dilution of the sample, the actual protein concentration of the sample to be measured.

5. In addition: if the spectrophotometer is used to determine, the glass test tube or glass colorimetric tube is used as the reaction vessel to make standard curves. After proper dilution of the protein sample to be tested, add it to a new glass test tube or colorimetric tube with a sample size of 1 mL. Add 3 mL of Coomassie brilliant blue G250 solution to the standard curve gradient tube and sample tube, mix thoroughly, stand at room temperature for 3-5 min, then use spectrophotometer for colorimetric detection.

6. Detection: The wavelength of the spectrophotometer is set to 595 nm, and the standard curve and the sample to be tested are zeroed with the standard curve No. 0 tube as the reference. Standard curves are drawn as described in Step 4 and protein concentrations in the samples to be tested are calculated.

Note

1.  The product should be restored to room temperature before use, and mixed upside down that avoid to affect the sensitivity of detection.

2.  For your safety and health, please wear safety glasses, gloves, or protective clothing.

For Research Use Only!

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