Strong 5'AMP-sepharose 4B (?)

[ssultang]

Hi, Everyone!

I am purifying some aldehyde dehydrogenases requiring NAD with
5'AMP-sepharose 4B.
But,I could not find the activity after 5'AMP-sepharose 4B affinity
chromatography

In column work,
    - starting and washing buffer : 10mM sodium phosphate buffer
    - eluting bnuffer : containing 0.5, 3, 7,10mM NAD
    - and at last eluting with 100mM sodium phosphate buffer.
Where is the enzyme?
How can I detach the guys from 5'AMP-sepharose 4B?

Please send me your idea.
With best regards.
Thank you in advance for your reply......