I would like to quantify the total amount of protein I have in red blood cells lysate samples to be able to load a precise amount in a western blot.
The red blood cells are lysate in a RIPA buffer but I could use another lysis buffer.
So far I tried BCA assay but the hemolglobin absorbes light in the same range so it gives me always the same value even with differents dilutions and I tried Nanodrop this afternoon and it gives me negatives value, even with a RIPA blank.
I'm looking for an assay I can use that doesn't interfere with the red color and the hemoglobin.
I have the same problem with the cytosolic fraction of my red blood cells because it is also red.
Any help is welcome.
Thank you in advance!
This paper (https://pubs.acs.org/doi/full/10.1021/acs.jproteome.7b00025#_i2)
used "a tryptophan fluorescence-based WF assay in microtiter plate format" and gave a reference for the method:Article Fast and Sensitive Total Protein and Peptide Assays for Prot...
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Liquid chromatography-mass spectrometry (LC–MS) analysis on a Q-Exactive Orbitrap instrument could be used to profile proteins. The details of procedure could be found in this articles: Article A robust mass spectrometry method for rapid profiling of ery...
Article Absolute proteome quantification of highly purified populati...
Qubit uses a fluorescence based approach and much more accurate than nanodrop if you can get your hand onto one.
Thank you Maurice Ekpenyong Navid J. Ayon Adam B Shapiro for your answers!
Adam B Shapiro this is exactly what I'm looking for!
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