Protocols.io provides an interactive version of this protocol where you can discover and share optimizations with the research community. Protein assays are routinely used in many research fields to estimate proteins in a vast array of buffers and conditions. A major problem for researchers is to select a protein assay from the vast selection on the market that is compatible with their protein sample. CB-X™ Protein Assay eliminates this problem as it is designed to be compatible with all commonly used buffers and conditions in protein isolation, storage and assays.
For protein samples in simple, uncomplicated, aqueous buffers, CB-X™ is a highly sensitive, single reagent assay that can be performed in 5 minutes. CB-X™ Protein Assay uses a protein dye that is an improvement on the Bradford Coomassie dye reagent (Figure 1)
For complicated protein samples, CB-X™ Protein Assay is supplied with reagents to clean up the samples and remove all reagents and chemicals that interfere with accurate protein estimation (figure 1). These reagents include detergents, chaotropes, reducing agents, alkylating agents, sugars, high salt concentrations, buffering agents and chelating agents (Table 1). The clean up stage and subsequent protein assay is performed in a single tube to ensure no protein loss and to maintain the accuracy of the assay. Figure 2 demonstrates the effect of CB-X™ in the presence of 1% Triton® X-100 or 1% SDS.
If the protein sample does not contain interfering agents, then a straightforward, single reagent assay is performed to give a linear response. If interfering agents are present or if artifactual results are produced, then the protein samples are treated with the clean up reagents and the protein is assayed to generate a linear response.
CB-X™ Protein Assay is supplied with lot-specific CB-X™ Tables. These allow researchers to perform single protein clean ups, subsequent assays, and then look up their absorbance in the CB-X™ Table to find the protein concentration. The CB-X™ Table eliminates the need for multiple protein stand ards and saves considerable time and effort. The table is prepared with a complex protein mixture that compares well with proteins from mammalian, plant, bacteria and yeast sources.The assay is supplied with a BSAprotein stand ard or anon animal protein stand ard with Cat. # 786-12X or 786-894 respectively for generating curves when using CB-X™ Assay Dye alone or for researcher’s who prefer to generate their own stand ard curve or their own CB-X™ Table per their specific conditions.
The CB-X™ Protein Assay is reliable over the range of 0.5-50µg per assay. The regular size kit contains enough CB-X™ Assay Dye for 500 protein assays and enough clean up reagents for 250 clean ups.
(Patents Pending) Table 1: CB-X™ Protein Assay is compatible with many interfering agentsDETERGENTS | REDUCING AGENTS |
Brij® 35 | 2% | 2-mercaptoethanol | 1M |
CHAPS | 2% | DTT | 1M |
CHAPSO | 2% | CHAOTROPES |
Nonidet® P-40 | 2% | Guanidine.HCl | 6M |
SDS | 2% | Urea | 6M |
Triton® X-100 | 2% | SALTS |
Tween® 20 | 2% | Ammonium sulfate | 1M |
Deoxycholate | 0.1% | MISCELLANEOUS |
SUGARS | EDTA | 0.1M |
Glucose | 1M | HEPES | 0.1M |
Sucrose | 25% | MES | 0.1M |
Features
- Unique cleaning step removes all interfering agents, including detergents and chaotropes, to improve accuracy
- Clean up and assay performed in a single tube to ensure NO protein loss
- Sensitive: 0.5-50µg linear response
- High reliability and reproducibility
- Long shelf life- stable for 12 months
Applications
- An improved, sensitive Bradford assay for most samples, including problematic samples, that is compatible with ALL common laboratory agents
- CB-X™ has been used in a wide array of techniques (Ref. 1-6) and applications including:
- Protein estimation in protein purification, electrophoresis, immunoanalysis, cell biology, molecular biology and other research applications
- Protein samples containing common laboratory agents, detergent solubilized membrane proteins and dilute protein solutions