Hi, Dear All,

I have a problem with SEC column when detect hGH (human growth hormone).

After 400 injections, peak of hGH was still perfect (TP=9000, asymmetric factor=1.1). Then, I stored it in 25% ACN after rinsing thoroughly with RO water.

But, when I used it 2 weeks later, serious tailing appeared (TP=900, asymmetric factor=2.4).

So, I rinsed the column with R.O. water --> 0.5N NaCL (pH3.0) --> RO water --> 0.1% SDS --> RO water --> 25% ACN --> 75% ACN --> 100% ACN --> 75% ACN --> 25% ACN --> RO water.

After that, TP=2500, asymmetric factor=.7~1.2. But, it couldn't hold long, now leading seems to appear.

My question is:

1) Why this could happen? I suppose if there was any hGH left in column, it could have been denatured by 25%ACN during storing period. Is that right?

2) What should I do further to recover the column?

Thanks! Look forward to hearing from you.

PS: Mobile phase is: 97% 0.05M PBS + 3% Propanol, pH 7.0


Wen Qu Ph.D.
University of Tennessee

What column do you use?