[3dem] Unrecognised CryoEM contaminant 2
rkhayat at ccny.cuny.edu
rkhayat at ccny.cuny.edu
Fri Mar 28 12:13:50 PDT 2014
There is the possibility that freezing causes the particles to
explode, leaving the condensed RNA/DNA behind. I've had VLPs
explode due to the freezing process when making cryo-EM grids.
Can you flash freeze the virus? Try flash freezing the virus,
thawing it, and checking how it affects your particles via
negative stained EM. This is probably the easiest way to test
if it might be the freezing.
If it's something in your sample, its presence should decrease
as you decrease the concentration of your particles.
Reza Khayat, PhD
Assistant Professor
The City College of New York
Department of Chemistry, MR-1135
160 Convent Avenue
New York, NY 10031
Tel. (212) 650-6070
---- Original message ----
>Date: Fri, 28 Mar 2014 17:04:21 +0000
>From: 3dem-bounces at ncmir.ucsd.edu (on behalf of "Hampton,
Cheri" <cheri.m.hampton at emory.edu>)
>Subject: Re: [3dem] Unrecognised CryoEM contaminant 2
>To: Daniel Luque <dluque at cnb.csic.es>
>Cc: "3dem at ncmir.ucsd.edu" <3dem at ncmir.ucsd.edu>
>
> Hi, I wonder could it be something in your buffer?
> It surrounds the spheres rather than laying on top.
> If the sample is dry from evaporation rather than
> blotting, strange things can precipitate.
>
> Cheri Hampton, PhD
> Emory University
> 914-413-8809
> On Mar 28, 2014, at 8:53 AM, "Daniel Luque"
> <dluque at cnb.csic.es> wrote:
>
> Hello
>
>
>
> Before a couple of days waiting for the list
> moderator approval to attach the images in the
> list , I do not have received any response (either
> positive or negative) and I do not have find a
> mail/way for contact. Thus, I have made the images
> available at our server at:
>
>
>
> http://halley.cnb.csic.es/~virus/3DEM/Example_images.pdf
>
>
>
> Bellow is text of the original question:
>
>
>
> We are doing CryoEM of viral particles. When we
> check the sample by negative staining, a field of
> intact homogeneous particles without significant
> contaminants is observed. However, when we
> visualize the same sample by CryoEM, additionally
> to a lower-than expected number of particles, we
> observe an unexpected contaminant mostly located
> in the holes (see attached document with some
> example images). Does anybody see this class of
> contaminants? Any idea about its origin?
>
>
>
> Thanks in advance!
>
>
>
>
>
> Daniel Luque
>
>
>
>
>
>
>
>
>
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