[3dem] Seeking Advice on BSL-2 CryoEM Laboratory Setup and Safety Protocols

Thu Jun 12 06:38:38 PDT 2025

Dear Matthijn and Kim,
Thank you both for your thoughtful responses and for sharing your valuable experiences.
Matthijn, I appreciate your clarification on the physics of cryo-TEM conditions. I apologize for not doing enough research before raising my initial question. As I understand now, under standard cryo-EM doses (such as 20–60 e⁻/Å² for single particle analysis or 100–120 e⁻/Å² for tomography), the risk is extremely low. However, I am curious about what might happen at much higher doses—on the order of thousands of e⁻/Å². Would such extreme irradiation potentially change the situation? From my current understanding, cryo-TEM operates far below both the triple point pressure and temperature of water, placing the sample firmly in the solid or vapor region of the phase diagram, where aerosol formation is not possible. Thus, the question is whether there might be any potential risks associated with the vapor that is formed under these conditions.
Additionally, I am interested in situations where the sample might inadvertently warm up in the TEM, for example, due to insufficient liquid nitrogen or a vacuum pump failure. Would there be any recommended follow-up procedures in such scenarios?
Kim, thank you for sharing your insights on the practical aspects of equipment disposal. It is certainly an important concern, and we will communicate with our local TFS engineer regarding this. Fortunately, our vacuum pumps have been quite stable over the past half year, and no replacement has been required so far.
Thank you both again for your guidance and support.
Best regards,
Jason
________________________________
From: Gibson, Kimberley <kimberley.gibson at yale.edu>
Sent: Thursday, June 12, 2025 21:04
To: Jason NG (CPOS) <jasonng.cpos at hku.hk>; Matthijn VOS <matthijn.vos at pasteur.fr>
Cc: 3dem at ncmir.ucsd.edu <3dem at ncmir.ucsd.edu>
Subject: Re: [3dem] Seeking Advice on BSL-2 CryoEM Laboratory Setup and Safety Protocols

Hi Jason,

Typically, at BSL-2, all equipment can be operated under normal conditions - i.e. in the open on a lab bench (Vitrobot or Leica  GPS) with special considerations for the evacuation of N2 gas or Ethane and Ethane/Propane mixtures.

Issues regarding BSL-2 have only arisen during disposal of equipment. TFS was not allowed to ship an old Ion Getter Pump overseas (from the US back to Eindhoven, NL) if the microscope operated in a BLS-2 lab. The IGP had to be disposed of locally and typically sprayed down with Ethanol prior to disposal. I have also sprayed an old Peltier device removed from a Vitrobot and sent it for electronic waste disposal.

Best,

Kim

________________________________
From: 3dem <3dem-bounces at ncmir.ucsd.edu> on behalf of Matthijn VOS via 3dem <3dem at ncmir.ucsd.edu>
Sent: Thursday, June 12, 2025 06:32
To: Jason NG <jasonng.cpos at hku.hk>
Cc: 3dem at ncmir.ucsd.edu <3dem at ncmir.ucsd.edu>
Subject: Re: [3dem] Seeking Advice on BSL-2 CryoEM Laboratory Setup and Safety Protocols

Dear Jason,

could you explain the physics hiw sn electron beam can create aerosols on a solid sample ar -196 degrees in ultra high vacuum (orders of magnitude under the tripple point of water)

I don’t see how this is possible.

cheers

Matthijn

On Jun 12, 2025, at 11:01, Jason NG (CPOS) via 3dem <3dem at ncmir.ucsd.edu> wrote:

Dear CryoEM Community,

We are currently evaluating the risks of becoming BSL-2 CryoEM laboratory and would greatly appreciate your guidance on several safety and setup considerations.

Specifically, we are interested in understanding:

  *   Given that the electron beam can potentially generate aerosols or cause warming within TEMs, what disinfection protocols or measures have you found effective for maintaining biosafety?
  *   Is implementing negative room pressure necessary in a CryoEM lab at BSL-2?
  *   For sample preparation instruments such as the TFS Vitrobot and Leica GP2, is it recommended to place them inside a biosafety cabinet? How about large instruments like the Leica ICE—are there specific safety measures we should consider?

Your insights and experiences would be extremely helpful as we evaluate the best practices for creating a safe and effective CryoEM environment. Thank you very much for your time and guidance.

Best Regards,

Jason NG

Senior Technical Assistant

LKS Cryo-EM Laboratory

<image001.jpg>

Centre for PanorOmic Sciences

LLG03, Laboratory Block

21 Sassoon Road, Pokfulam, Hong Kong

P: +852 3910-3528

E: jasonng.cpos at hku.hk<mailto:jasonng.cpos at hku.hk>

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