[3dem] Unrecognised CryoEM contaminant 2

[rkhayat at ccny.cuny.edu]

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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