[3dem] Ethane ice on Vitrobot frozen grids

Sacha De Carlo sachadecarlo at yahoo.com
Fri Dec 12 12:41:04 PST 2008


There are many reasons that lead to a lot of ethane on your grid to a point where it becomes unusable to the point you're describing.

Have you tried some of the following:
- monitor the temp. with a thermocouple
- wait that the ethane is at the right temperature, visually monitor the formation of solidified ethane at the bottom of the aluminum cup, then add some fresh ethane and only then plunge (before it solidifies again)
- if some ethane is on the grid (if you can see it, that is), flush it away by pouring some liquid nitrogen on to the grid (clipped securely on the tip of the cryo-holder)
- remove it delicately with tweezers (from the side, do not punch a hole from above!)
- keep your tweezers wiped clean and dry in between preparing new grids
- last but not least, try not to be too sloppy (meaning, keep the ethane cup clean, do not flush nitrogen in there)

Good luck!

S.


================================================
Sacha De Carlo, Assistant Professor
Chemistry Department, Marshak Science Building &
New York Structural Biology Center
The City College of New York
Convent Ave & 138th Street
New York, NY 10031
(212) 650-6070
e-mail: sdecarlo at ccny.cuny.edu
URL: http://www.planetesacha.com
================================================

--- On Fri, 12/12/08, Paul Keller <kellerp at mail.nih.gov> wrote:
From: Paul Keller <kellerp at mail.nih.gov>
Subject: [3dem] Ethane ice on Vitrobot frozen grids
To: 3dem at ncmir.ucsd.edu
Date: Friday, December 12, 2008, 3:27 PM

Hello All,

I was wondering if anyone on the list had any insight into something I observe
regularly when using our Vitrobot (Mk II); the formation of a layer of ethane
ice on the grid during transfer from the liquid ethane cup into the outer ring
and grid storage box.  The grid cannot then be used until the ethane ice is
removed.

The ethane layer almost always seems to dissipate if I leave the grid in LN2
storage at least overnight, but this is not always desirable.  Does anyone on
the list have any advice for removing the ethane ice without damaging the
underlying sample, or preferably, techniques for avoiding the formation of an
ethane ice layer entirely?

Thank you,

-Paul Keller
Laboratory of Structural Biology Research
NIAMS, NIH
Building 50, Room 1504
50 South Drive MSC 8025
Bethesda, MD 20892-8025
301-451-4939
kellerp at mail.nih.gov

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