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Commassie Plus Protein Assay Kit

AR0145

1 kit

20 USD

AR0145

Commassie Plus Protein Assay Kit

1 kit

20 USD

Western Blotting Related Reagent

4˚C for one year. (BSA Standard should be stored at -20˚C).

The Coomassie plus protein assay kit is a quick coomassie-binding, colorimetric method for total protein quantitation. When coomassie dye protein in an acidic medium, an immediate shift in absorption maximum occurs from 465 nm to 595 nm with a concomitant color change from brown to blue. Performing the assay in either test tube or miscroplate format is simple：Combine a small amount of protein sample with the essay reagent, mix well, incubate briefly and measure the absorbance at 595 nm. Protein concentrations are estimated by reference to absorbances obtained for a series of standard protein dilutions, which are assayed alongside the unknown sample. When protein combined with Commassie, they will arrive at balance within 2-5 min, the complete reaction is very quick, and the combo could maintain stability for 1 hour at room temperature whose sensibility is as 4 times as Lowry method.This kit can measure microgram of protein content and the test range of protein concentrations is 0～1500ug/ml

Ⅰ. Test Tube Procedures A. Standard Test Tube Procedure (Working Range = 100-1,500 ug/ml) Note: The linear working range of BSA = 125-1,000 ug/ml; the linear working range of IgG = 125-1,500 ug/ml. 1. Pipette 0.05 ml of each standard and unknown sample into appropriately labeled test tubes. 2. Add 1.5 ml of the Coomassie Plus Reagent to each tube and mix well. 3. Optional: For the most consistent results, incubate samples for 10 minutes at room temperature (RT). 4. With the spectrophotometer set to 595 nm, zero the instrument on a cuvette filled only with water. Subsequently, measure the absorbance of all the samples. 5. Subtract the average 595 nm measurements for the Blank replicates from the 595 nm measurements of all other individual standard and unknown sample replicates. 6. Prepare a standard curve by plotting the average Blank-corrected 595 nm measurement for each BSA standard vs. its concentration in ug/ml. Use the standard curve to determine the protein concentration of each unknown sample. B. Micro Test Tube Protocol (Working Range = 1-25 ug/ml) 1. Pipette 1.0 ml of each standard and unknown sample into appropriately labeled test tubes. 2. Add 1.0 ml of the Coomassie Plus Reagent to each tube and mix well. 3. Optional: For the most consistent results, incubate samples for 10 minutes at room temperature (RT). 4. With the spectrophotometer set to 595 nm, zero the instrument on a cuvette filled only with water. Subsequently, measure the absorbance of all the samples. 5. Subtract the average 595 nm measurement for the Blank replicates from the 595 nm measurements of all other individual standard and unknown sample replicates. 6. Prepare a standard curve by plotting the average Blank-corrected 595 nm measurement for each BSA standard vs. its concentration in ug/ml. Use the standard curve to determine the protein concentration of each unknown sample. Ⅱ.Microplate Procedures A. Standard Microplate Protocol (Working Range = 100-1,500 ug/ml) 1. Pipette 10ul of each standard and unknown sample into the appropriate microplate wells. 2. Add 300 uml of the Coomassie Plus Reagent to each well and mix with plate shaker for 30 seconds. 3. Remove plate from shaker. For the most consistent results, incubate plate for 10 minutes at room temperature (RT). 4. Measure the absorbance at or near 595 nm with a plate reader. 5. Subtract the average 595 nm measurements for the Blank replicates from the 595 nm measurements of all other individual standard and unknown sample replicates. 6. Prepare a standard curve by plotting the average Blank-corrected 595 nm measurement for each BSA standard vs. its concentration in ug/ml. Use the standard curve to determine the protein concentration of each unknown sample. B. Micro Microplate Protocol (Working Range = 1-25 ug/ml) 1. Pipette 150 ul of each standard and unknown sample into the appropriate microplate wells. 2. Add 150 ul of the Coomassie Plus Reagent to each well and mix with plate shaker for 30 seconds. 3. Remove plate from shaker. For the most consistent results, incubate plate for 10 minutes at room temperature (RT). 4. Measure the absorbance at or near 595 nm with a plate reader. 5. Subtract the average 595 nm measurements for the Blank replicates from the 595 nm measurements of all other individual standard and unknown sample replicates. 6. Prepare a standard curve by plotting the average Blank-corrected 595 nm measurement for each BSA standard vs. its concentration in ug/ml. Use the standard curve to determine the protein concentration of each unknown sample. Note: 1. When Pipette sample should assure the greatest accuracy. 2. The reagent added with sample and the solution in each tube should be placed still to react after turned upside down and mixed and equilibrated. Avoid shake again during measurement. 3. For greatest accuracy, after adding reagent, please measure the absorbance in 5～20 min, because the color is most stable in this period. 4. Some positive ion, such as K+. Na+. Mg2+. （NH4）2SO4 and ethyl alcohol would not interfere the measurement, but a plenty of decontaminating agents like TritonX-100. SDS would do interfering substances.

For quantitation of total protein

Tube procedure: 50 assays Microplate procedure: 300 assays