Among all different protein content assays (BCA, Lowry, Bradford, etc) which one would you recommend for under microgram amounts of protein mixtures?
Hi Reza
I would go for BCA which has sensitivity range of 0.5-10 micro grams/ml
Good luck
I would like to recommend HPLC-protein assay method; i.e., HPLC-protein assay method shows ca. 200-fold higher sensitivity than Lowry's or other photometric methods (photometric methods gives false results due to using non-linear calibration-curve). HPLC-protein assay uses reliable linear-calibration line inserting to the zero point. This is the most impotant point; i.e., ELISA or RIA is not able to truly determine the protein amount due to the absence of this type of linear-calibration line.
HPLC-protein assay uses the detection at 210nm (measures peptide bonds),and sensitivity increases than using 254 nm or 280 nm (measures absorption of Tyr, Phe, Trp). Use of 210 nm is only possible by separation with HPLC. Further, possible interference due to polysaccharides (containing GlcNAc, GalNAc, NANA) is not observable by using real sample specimens such as serum, milk, bile, pancreatic juice, tissue homogenates show no interference on the assay (see file HPSEC protein...); i.e., UV absorption at 210 nm is high in proteins as compared to in polysaccharides.
Further, it is noteworthy that fluorometric assay is only to prevent the interferences by other molecules; i.e., fruorometric photometry increases specificity, but the level of sensitivity is similar to photometry.
Here are some reagents for very sensitive fluorescence-based protein quantitation.
https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-assays-analysis/protein-assays/fluorescent-protein-assays.html
Dear Dr. Hayakawa and Dr. Shapiro thank you very much for your enlightening comments.
- Badges
- Science topic
- Similar topics
- Microbiology
- Assays