I want to know the concentration/activity of alkaline phosphatase (ACP) in an animal tissue (liver) from the absorbance. I have the following data with me;
tissue weight, volume of supernatant used as test sample, absorbance and standard curve of the standard used.
All the reagents were prepared in the lab and no kit was used for the test.
Alkaline phosphatase produces P from p-nitrophenyl phosphate (pNpp). I assume that your cuvette is 1 cm in length; if so, then subtract absorbance of blank from that at 405 nm. Convert this difference into phosphate produced applying the Beer-Lambert's equation; difference = c * Epsilon * length
c = difference/(1 cm * 18300 M-1 cm-1 *) = moles/liter
Now take into account the volume of your cuvette, is it 3 ml? Then multiplying per 0.003 mL = moles in 15 min, i.e.
in 1 min = mol/min = Convert to nmol/min = Units
The enzyme activity in the extract can be expressed as specific activity, µmoles of product formed per minute per mg of protein. So you need to do a protein assay on the extract.
- Badges
- Science topic
- Similar topics
- Biological Science
- Anatomy
- Tissue