Conversion of milligram weighing Alkaline phosphatase powder to U/L solution - (Jan/27/2016 )

Hi all,

I want to detect serum enzyme Alkaline phosphatase using colorimetric assay(BCIP/NBT). For that I need to plot a calibration curve, in which the X axis should be in U/L of Alkaline phosphatase (ALP). I have only ALP in the form of powder weighing 100mg. My question is how to convert the mg weighing ALP to U/L solution so that I need to obtain solutions in the range of 1U/L, 2U/L,5U/L,10U/L, 100U/L, 1000U/L.

bcip/nbt produces an insoluble colored product. this is good if you are using it to develop a western blot or determining activity in a gel. if, however, you are trying to look at a result in solution then you would be better off using para-nitrophenyl phosphate as your substrate.

as for your concentration question, the data sheet that came with the alkaline phosphate should give you the U/mg protein (or mg solid, not necessarily the same).

Thank u for the reply, min. of 20 DEA Units/ mg protein is given in the datasheet of ALP. Using this, how can I convert U/mg to U/L? Can you give me any references regarding this?

Thanks in advance

did you receive a lot analysis sheet? if not then you may be able to download from the supplier's website.

The lot analysis will tell you how many units/mg are in the batch you received.

if you want to prepare a specific concentration (u/l) then just add the amount (mg) of enzyme that gives you the number of unite you require for the volume you are preparing.

if you want to determine the u/l concentration in the vial of suspended protein then you determine the number of total units by multiplying u/mg by total mg. you then divide total units by the volume (in liters) in the vial.