Hi,
someone can help me?
regarding problems that occurred regarding the following two columns:
- DNAPac™ PA200, P/N: 63000, S/N: 007957, L/N: 2101045
- DNAPac™ PA200 RS, P/N: 82509, S/N: 007957, L/N: 2101045
We are using those columns on a HPLC for method development to analyze siRNA (double stranded, approx. 20-mers, chemically-stabilized oligonucleotides) under denaturing conditions (T=80°C).
We’ve been testing a great variety of different buffer systems having no issues with the columns stated above.
Lately however, we observed increases back pressure from those columns significantly above the values stated in the Troubleshooting Guide:
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The following solvents we were using while this problem occurred:
- Solvent A: 20 mM Na2HPO4, 10% ACN in water (pH 9.5)
- Solvent B: 1 M NaCl, 20 mM Na2HPO4, 10% ACN in water (pH 9.5)
Extensive column cleaning as described in the manual didn’t bring any improvement.
Additionally, we observed an continuous increase of pressure while running successive injections before the pressure is now too high to run any analysis within the pressure limits of the columns.
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Is there anything obvious to you we did wrong to have caused the columns to clog? What are potential explanations for such a rapid back pressure increase that can not be reduced by extensive cleaning? How could we make sure that this will not happen again in case we ordered new columns because besides this problem we are quite happy with the performance of the columns.
I hope we can figure something out.
Please let me know in case you have questions or need additional information.
Thank you!
Best regards,