TMB in ELISA at 630nm - (Nov/11/2002 )
using an TMB substrate in an ELISE experiment requires me to measure at 655 nm, but our old MRX microplate reader can only read at 630nm. Off my head, i would not think that it would prove a huge problem, but just in case..... Measuring it at 450 nm could also be an opportunity, but it requires an additional step that i I might now want to introduce in the experiment.
The problem is that the company, Dynatech, who produced the filters for the microlpate reader does not, to my knowledge, exist any longer.
But the problem: is it completely stupid to measure at 630 nm, or should I buy a new apparatus, like the Biorad model 550?
I use Tetramethlybenzidine (TMB) in ELISA myself, and I've the same problem, that is our ELISA reader can't do the wavelength. Better said, I don't have the same problem anymore, since I stop the reaction with 2 N H2SO4 and read the plates at 450nm. Works very fine for me & isn't too complicated.
We use ELSIA kit from Pharmingen now and they even put the stopping step in their protocols...
Never give up, never surrender!