Nuclear Lysates Protein Quantification - Nuclear Extracts (Sep/20/2007 )
Wondering if someone could help me out here. I prepared some nuclear lysates to run a trans-binding ELISA. I measured the protein using the QUANT-IT kit by Probes. The ELISA ran pretty smoothly. Well I had to run it a second time with more tissue, but this time I measured the protein using the common BCA assay, since i was out of QUANT-IT.
My concenrn is, that I saw much higher values for protein concentrations when I measured it using BCA. Which one do I believe 
?
HELP..
homogenize well samples. then do each point 2 times.
Your blank is ok, and did you make the reference points ?
Your blank is ok, and did you make the reference points ?
Homeogenized the well. Ran them in triplicates, but still off. Yes, I have standards also. Im worried that the nuclear lysis buffer has some glycerol in it (Nuclear Kit from Panomics), which may be interfering.
i guess so.
I mean, we use platic cuvettes.
Ae my lab is low fund, we tend to reuse them by rinsing with ddH20.
Once, i rinsed after with EtOH, and saw a tigh blue color.
So i guess alcohol turne bradford little blue... and guess so do the glycerol
i realize at this time, i've never do the test to put little amount of glycerol only to see if it turns the coloration.
I mean, we use platic cuvettes.
Ae my lab is low fund, we tend to reuse them by rinsing with ddH20.
Once, i rinsed after with EtOH, and saw a tigh blue color.
So i guess alcohol turne bradford little blue... and guess so do the glycerol
i realize at this time, i've never do the test to put little amount of glycerol only to see if it turns the coloration.
when we use plastic cuvettes for bradford, we wash them with meoh or etoh to clean off coomassie stuck to the surface.