[3dem] your opinion on these images from graphene grids

[David Michael Belnap]

Jacopo,

Assuming these are pictures of a frozen specimen, I would assume the spots are artifacts of likely radiation damage, especially if they bear no resemblance to your protein prepared by negative staining.  The spots also vary in size.  Does that resemble your protein?  If so, then you have a very heterogeneous sample.

David


> On Nov 20, 2019, at 6:45 AM, David Michael Belnap <David.Belnap at utah.edu> wrote:
> 
> Jacopo,
> 
> Are these images of blank grids or did you freeze a sample on the grid?
> 
> David
> 
> 
>> On Nov 20, 2019, at 6:40 AM, Jacopo Marino <jacopo.marino at psi.ch> wrote:
>> 
>> Dear Community,
>> 
>> I would like to ask you what you think about these images we took by using graphene grids (suspended monolayer graphene on TEM grid substrate, quantifoil gold, purchased from Sigma). We are puzzled because if one area is exposed with the usual 20 electrons for 2 seconds, it is very difficult to see if there are particles or not. However, when the same area is re-exposed with the same dose or for longer, there are particles-like entities appearing, which we wonder whether they could be an artefact from the longer exposure. Interestingly, by exposing for even longer (200 electrons), particles seem not be destroyed, which makes us doubting this is protein.
>> 
>> Here is the link with the images, where 001.tif is the image we took with normal exposure, 18.tif is exposed twice, while 19.tif is the one shot at least three times. We used a Talos at 200 kV, pixel size 1.61.
>> 
>> https://www.dropbox.com/sh/5rb59jwsiu6zcrq/AAAze6dKhR4P4Wzb7LMCs75Ba?dl=0
>> 
>> Thank you for your help !
>> 
>> Jacopo
>> 
>> -- 
>> Dr. Jacopo Marino
>> Laboratory of Biomolecular Research
>> Paul Scherrer Institute
>> OFLB/005
>> 5232 Villigen PSI, Switzerland
>> tel: +41 56 310 5484 (or +41 56 310 5777)
>> e-mail: jacopo.marino at psi.ch
>> 
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