Navigation Links
Total Cellular Protein Determination Using the DC Protein Assay


Introduction
The DC Protein Assay reagents contain both detergent and base which are required for cell lysis of many cell samples prior to determination of total cellular protein. The following protocol can be used for tissue cells; dry cells; protein precipitates from heat, acid, or acetone treatment; or any other samples that require both sodium hydroxide and detergent for solubilization.


Protocol
1. Wash cells or protein precipitate. Using an isotonic solution such as TBS or PBS, suspend the sample in excess buffer and mix by inversion or vortex mixer. Centrifuge sample or precipitate until a tight pellet has formed. Discard the supernatant.

2. Prepare working reagent A' by mixing 20 l Reagent S to each milliliter of Reagent A that will be needed for the assay. Mix by gentle inversion to prevent foaming.

3. Add 1 ml working reagent A' to approximately 1 x 106 cells. Vortex intermittently for 510 minutes, and heat to 60 C (if necessary) to solubilize the pellet.

4. Cool mixture to room temperature. In order to accurately determine protein concentration, several dilutions of the digestion with reagent A' should be made (1:2, 1:4, 1:8, etc.). Record the dilution factors for correction in the final protein concentrations.

5. Create a standard curve. Transfer 100 l of the standard into a clean test tube. Three to five data points should be generated. Either standard supplied with the kit (BSA or IgG) can be used, since both react similarly to the DC Protein Assay reagents.

6. Transfer 500 l sample of serially diluted samples from step 5 into clean test tubes.* To correct for volume difference between samples and standard curve, add 100 l water to each tube. Add 4.0 ml Reagent B and vortex.

7. Add 500 l working reagent A' to samples in the standard curve series and vortex.

8. Add 4.0 ml Reagent B, and vortex.

9. Incubate standards and samples at room temperature for 15 minutes. Measure absorbance at 650750 nm. Compare results to a standard curve.


For technical assistance, call your local Bio-Rad representative, or in the U.S., call 1-800-4BIORAD.


back to top
'"/>

Source:


Page: All 1 2

Related biology technology :

1. New Kit Generates Exceptionally Pure Total RNA for RT-PCR
2. High-Throughput Isolation of Total RNA
3. Isolate and Analyze Total RNA from Cells Harvested by Laser Capture Microdissection
4. Fast Isolation of Total RNA from Small Numbers of Cells
5. Total RNA Isolation Protocol 1,2,3
6. Isolate It All: siRNA miRNA Total RNA Native Protein
7. Isolate Total RNA and Protein From the Same Sample
8. Isolate High Quality Total RNA from LCM Samples Suitable for Microarray and qRT-PCR Analysis
9. Recover High Yields of Total Nucleic Acid from Formalin-fixed, Paraffin-embedded (FFPE) Tissue
10. Total RNA from Whole Blood for Expression Profiling
11. RNAPrep - Total RNA Isolation Purification
Post Your Comments:
*Name:
*Comment:
*Email:


(Date:10/10/2017)... (PRWEB) , ... October 10, ... ... development-stage cancer-focused pharmaceutical company advancing targeted antibody-drug conjugate (ADC) therapeutics, today confirmed ... targeted HPLN (Hybrid Polymerized Liposomal Nanoparticle), a technology developed in collaboration with ...
(Date:10/10/2017)... , Oct. 10, 2017 SomaGenics announced the ... NIH to develop RealSeq®-SC (Single Cell), expected to be ... small RNAs (including microRNAs) from single cells using NGS ... the need to accelerate development of approaches to analyze ... "New techniques for measuring levels of mRNAs ...
(Date:10/9/2017)... ... October 09, 2017 , ... The award-winning American Farmer television ... quarter 2018. American Farmer airs Tuesdays at 8:30aET on RFD-TV. , With global ... the challenge of how to continue to feed a growing nation. At the same ...
(Date:10/7/2017)... ... October 06, 2017 , ... ... consulting for microscopy and surface analysis, Nanoscience Instruments is now expanding into ... a broad range of contract analysis services for advanced applications. Services will ...
Breaking Biology Technology:
(Date:4/19/2017)... York , April 19, 2017 ... as its vendor landscape is marked by the presence ... market is however held by five major players - ... Together these companies accounted for nearly 61% of the ... the leading companies in the global military biometrics market ...
(Date:4/13/2017)... 2017 UBM,s Advanced Design and Manufacturing event ... emerging and evolving technology through its 3D Printing and ... alongside the expo portion of the event and feature ... focused on trending topics within 3D printing and smart ... event will take place June 13-15, 2017 at the Jacob ...
(Date:4/11/2017)... Research and Markets has announced the addition of the ... ... grow at a CAGR of 30.37% during the period 2017-2021. ... prepared based on an in-depth market analysis with inputs from industry ... over the coming years. The report also includes a discussion of ...
Breaking Biology News(10 mins):