I ran sephadex column for the plant protein and took absorbance at 280 nm. When I repeated the experiment it doubled the absorbance value for the same sample. Why is it so?
When you say you repeated the experiment do you mean you re-prepared the plant protein extract? May I also know how did you prepare the extract? Because the efficiency of extracting proteins from any sources might differ greatly, in a batch by batch basis.
Also, what type of proteins are you looking at? Enzymes?
Sounds like experimental error or batch to batch variation. Did you run the same sample twice over the column or did you make a new one?
Accurate A280 readings are done under denaturing conditions. Use Bradford or another assay as control if you believe the UV reading is altered and not the concentration.
I ran the same sample. Moreover, I have already prepared the extract once and stored in the form of aliquots. The thing I changed was only the aliquote. Please suggest me what to do? Thanku.............
- Badges
- Science topic
- Similar topics
- Social Psychology
- Attitude