CE - Urea creep

[biochemist]

How do you prevent urea creep?

Is it considered serious for an Agilent CE system, if the urea crystallizes all around the vial cap but doesn't appear on the outside of vials?

[dorota]

We have had problems with crystals on the vial caps. In our case it was because we have kept waste vials empty as recommended by an application guide (which you can read here: http://www.beckmancoulter.com/wsrportal ... A78788.pdf). Now we fill these vials with water and problems have gone. But I do not know if it is remedy for your system - we have Beckman CE.

[biochemist]

Thank you!

How much water did you fill in the waste vials?

Did the urea crystals ever interfere with your analysis prior to that?

[dorota]

About 0.5-1 ml. The end of capillary must reach water in the vial otherwise the drop of buffer formed at the end of a capillary may spread on the top of the next vial cap and after water evaporation buffer components may crystallize.

The answer for your question regarding the influence of these crystals on analysis is more complicated and I still do not know if our problems where connected with these crystals. We use CIEF to determine pI and to study conformational stability of proteins under different concentration of urea. It was a time when the system worked almost 24h/day 7days/week. Results did not arouse suspicion but during analysis the system accidentally was bursting capillaries. One end of a capillary punched a vial cap and consequently a capillary was broken. If I remember correctly 11 capillaries where demolished during analysis over 1 year! Service could not find any reason for this. During an inspection they found crystals on the caps and recommended not to use empty vials what was implemented in our procedure. Since then no single capillary was broken. But I must underline that we do not use CIEF so often as we did in the past. I can hardly believe that these crystals caused capillaries to punch vial caps but I do not have better explanation. I hope the future will bring the answer. One way or another I think it’s better not to have crystals on vial caps.

[cari]

As Dorota rightly remarked, some liquid in the waste vials is important, and not only for urea solutions.

The other thing which I encounter often when asked to come troubleshooting, is insufficient cleaning of the instrument. Whether you run Beckman, Agilent or another piece of equipment, please clean regularly It will safe you a lot of issues. With simple, highly soluble and non-viscous buffers, once a week is often sufficient. If the buffer contains gel, urea, high concentrations of e.g. cyclodextrins, or is viscous for other reasons: clean after every sequence.
Often rinsing with water suffices. If you have an Agilent instrument, don't forget the prepunchers.

Furthermore, don't overfill the vials (max 3/4 fill height, see also the vendors manual) and please don't reuse the vials and caps with solutions that are viscous, contain urea or anything in high concentration.

All together, this should reduce the urea creeping around.
/Cari
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