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Increasing the amount of phosphoric acid enhances the suitability of Bradford assay for proteomic research
- Source :
- ELECTROPHORESIS. 40:1107-1112
- Publication Year :
- 2019
- Publisher :
- Wiley, 2019.
-
Abstract
- The Bradford assay is one of the most commonly used methods for protein quantification. However, in proteomic research, the lysis buffer generally used for dissolving proteins can cause some interference to the assay. The dye reagent of classical Bradford assay contains 8.50% (w/v) phosphoric acid, which is an important factor relating to the color yield of the assay. In this study, the phosphoric acid content in dye reagent was increased to 9.35% (w/v), 10.20% (w/v), and 11.05% (w/v) to evaluate the changes of interference and the effects of lysis buffer on the interaction between proteins and dye reagent. Results show that lysis buffer not only causes background interference but also affects the protein-dye chromogenic process. Analysis of different components in the lysis buffer showed that carrier ampholyte is the main factor that introduces interference to the Bradford assay. Detergents are well-known interfering compounds in the Bradford assay, but CHAPS and octyl b-D-glucopyranoside only cause slight interference. When the amount of phosphoric acid was increased from 8.50%(w/v) to 10.20% (w/v), the sensitivity of the Bradford assay to proteins in lysis buffer was increased, and the interference delivered by lysis buffer was considerably reduced.
- Subjects :
- Proteomics
Ovalbumin
Detergents
Clinical Biochemistry
02 engineering and technology
Buffers
01 natural sciences
Biochemistry
Analytical Chemistry
chemistry.chemical_compound
Chaps
Two dimensional electrophoresis
Lysis buffer
Phosphoric Acids
Phosphoric acid
Bradford protein assay
Chromatography
Chemistry
Chromogenic
010401 analytical chemistry
Proteins
Globulins
Serum Albumin, Bovine
021001 nanoscience & nanotechnology
0104 chemical sciences
Reagent
Yield (chemistry)
Biological Assay
Indicators and Reagents
0210 nano-technology
Subjects
Details
- ISSN :
- 15222683 and 01730835
- Volume :
- 40
- Database :
- OpenAIRE
- Journal :
- ELECTROPHORESIS
- Accession number :
- edsair.doi.dedup.....6cdfffafe7a5582f3a819d5e1ac28b9f