NADH nightmare - (Mar/04/2006 )

Hello,

I am having trouble preparing my NADH solutions for spectrophotometric assays.
I prepared a 5mM NADH solutions in 0.01 NaOH buffer. But when I blanked on the NADH solution, the O.D. crazy went down.

Can anyone suggest what is going on and how to fix it?
Thanks.

valnet
valnet_mi@hotmail.com

It seems that you are simply removing the hydrogen from the NADH. NAD+ is not spectroscopically active at 340nm and this maybe why the absorbance readings are off.

Try making the soln in a buffer with a pKa close to you desired pH.

Good luck
K-