[3dem] 3dem Digest, Vol 143, Issue 2

Mon Jul 1 11:42:12 PDT 2019

Hi Cindy,
Try to dissolve plastic with THF (Tetrahydrofuran). The only important tip
is to move fast because it evaporates very quickly
Good look
Pedro

El lun, jul 1, 2019 11:37, <3dem-request at ncmir.ucsd.edu> escribió:

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> Today's Topics:
>
>    1. Re: Help! Plastic films on my holey grids are KILLING ME
>       (Lars-Anders Carlson)
>    2. Re: Help! Plastic films on my holey grids are KILLING ME
>       (Mariena Silvestry Ramos)
>
>
> ----------------------------------------------------------------------
>
> Message: 1
> Date: Mon, 1 Jul 2019 19:23:10 +0200
> From: Lars-Anders Carlson <lars-anders.carlson at umu.se>
> To: <3dem at ncmir.ucsd.edu>
> Subject: Re: [3dem] Help! Plastic films on my holey grids are KILLING
>         ME
> Message-ID: <7aedd66e-c28f-40a6-769c-d8db322c0ab4 at umu.se>
> Content-Type: text/plain; charset="windows-1252"; format=flowed
>
> Hi Cindi,
> Can you remove it by just plasma cleaning longer? That could be the
> reason why some of us typically don't see this film in the first place
> (another reason could of course be that we don't look as closely as you).
>
> QF grids used to have plastic deposits around the edge of the holes,
> which is where they would start bubbling at higher exposures (80-100
> e/?2), but I haven't seen much of this lately.
>
> I hope you find a solution since you seem really frustrated about it.
>
> Cheers,
> Lars
>
> On 2019-07-01 18:38, Schwartz, Cindi (NIH/NIAID) [E] wrote:
> > Hi all,
> >
> > We?ve been suffering with trying to get the plastic films off our
> > Quantifoil grids with very inconsistent results. We?ve also had film
> > issues on the c-flats, but my understanding is that the Quantifoils use
> > a plastic background and the c-flats use a carbon background. Please
> > correct me if I?m wrong on that one. We use both copper and gold grids
> > with the carbon substrate of various different patterns from R1/4 to
> > Multi-A with grids that we purchased 2 years ago to ones we?ve received
> > last month.
> >
> > The list of solvents we?ve tried that should dissolve plastic films:
> >
> > Ethyl acetate
> >
> > Chloroform
> >
> > Ethylene dichloride
> >
> > Trichlorobenzene
> >
> > The list of solvents to clean them up a bit:
> >
> > Acetone
> >
> > Ethanol
> >
> > 5% acetic acid
> >
> > We?ve tried sonication (a disaster)
> >
> > Submerging in drops
> >
> > Submerging in vast quantities of liquid
> >
> > With/without little stir bars while the grids are on a metal platform in
> > the solvent
> >
> > Time ranges from 15 min to overnight
> >
> > With/without a light glow discharge before cleaning
> >
> > We?ve put 10 nm colloidal gold fiducials on both sides of the grids
> > (like you?d use for plastic section tomography?where they stick to
> > Formvar just fine) after cleaning with various permutations from the
> > above lists and we see that the carbon has gold, and the ?holes? are
> > clean with no gold, only they aren?t clean! The colloidal gold does not
> > seem to stick to the plastic film because we see the film if we negative
> > stain apoferritin on these same grids or we see the films in the big
> > scope based on the ctf or the way the ice has blotted or how the films
> > have broken away and flipped onto the carbon. It?s rather obvious when
> > you also see true holes next to ?filmed? holes.
> >
> > Lastly, something will work for a subset of grids in a box, but later
> > on, grids from the same box (even the same SIDE of the box) don?t get
> > cleaned using the same protocol.
> >
> > All I can conclude is that whatever this plastic is, we should get
> > Quantifoil to stop using it because nothing seems to dissolve it and
> > it?s probably killing the planet somehow. If not, it?s certainly
> > detrimental to my mental health. Or better yet, Quantifoil should do the
> > research and either clean them better themselves or at least give us a
> > protocol to do so in the lab.
> >
> > Any tips, tricks, or surefire ways to clean grids would be great. And
> > please publish it because nothing pops up on Pubmed when I search for it.
> >
> > Thanks,
> >
> > Cindi L. Schwartz
> >
> > Electron Microscopist
> >
> > Rocky Mountain Labs/NIAID/NIH
> >
> > 903 South 4th Street
> >
> > Hamilton, MT ?59840
> >
> > 406-363-9228
> >
> > Cindi.Schwartz at NIH.gov <mailto:Cindi.Schwartz at NIH.gov>
> >
> > Description: Macintosh
> > HD:Users:schwartzcl:Desktop:large-green-recycle-symbol-hi.png Please
> > remember our Earth before printing this email Description: Macintosh
> > HD:Users:schwartzcl:Desktop:large-green-recycle-symbol-hi.png
> >
> > *************************************************************************
> >
> > The information in this e-mail and any of its attachments is
> > confidential and may contain sensitive information. It should not be
> > used by anyone who is not the original intended recipient. If you have
> > received this e-mail in error please inform the sender and delete it
> > from your mailbox or any other storage devices. National Institute of
> > Allergy and Infectious Diseases shall not accept liability for any
> > statements made that are sender's own and not expressly made on behalf
> > of the NIAID by one of its representatives.
> >
> > *************************************************************************
> >
> >
> > _______________________________________________
> > 3dem mailing list
> > 3dem at ncmir.ucsd.edu
> > https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem
> >
>
> --
> Lars-Anders Carlson
> Assistant Professor
> Dept of Medical Biochemistry and Biophysics
> Wallenberg Centre for Molecular Medicine
> Ume? University
> 901 87 Ume?, Sweden
> http://www.carlsonlab.se/
>
>
> ------------------------------
>
> Message: 2
> Date: Mon, 1 Jul 2019 17:37:35 +0000
> From: Mariena Silvestry Ramos <ms3289 at cornell.edu>
> To: "3dem at ncmir.ucsd.edu" <3dem at ncmir.ucsd.edu>
> Subject: Re: [3dem] Help! Plastic films on my holey grids are KILLING
>         ME
> Message-ID:
>         <
> BN3PR04MB2260CA5C45471010C90F810ECBF90 at BN3PR04MB2260.namprd04.prod.outlook.com
> >
>
> Content-Type: text/plain; charset="windows-1252"
>
> Hi Cindi,
>
>
> I'm so sorry. Years ago, in Phoebe's lab, we used to get the grids with
> just plastic, then C-coat them ourselves, then dissolve the plastic. And
> indeed, that plastic was a nightmare to remove.
>
>
> A few yrs ago I shared this email with the list:
> https://mail.ncmir.ucsd.edu/pipermail/3dem/2012-December/002760.html
>
>
> >From your email, it looks like everything you're doing is pretty much
> what we did too, but in our experience (if I remember correctly), the
> plastic did come off.
>
>
> The only thing I didn't see in your email was baking the grids. In
> Phoebe's lab at Vanderbilt, we put our grids on top of an aluminum disk,
> took them to a CM12, cranked up the power and left them overnight inside
> the microscope (our conditions were, a beam spread slightly inside the 4cm
> mark, spot size 1, all apertures out). I don't think this is done anymore,
> but in our case, it seemed to work. Dewight used to also mention how
> obnoxious and stubborn that plastic was, but I don't remember if he had an
> extra step to completely get rid of the plastic. We did 2x wash with each
> of the 4 solvents I mention in that email, and waited until the filter
> paper was dry before adding the next solvent down the list.
>
>
> Finally, IDK if it would be advisable, but for hydrocarbons left behind
> (this is on holders), we've used an orange peel cleaner (we use Zep, but
> any other orange-based compound should work similarly). If all else fails,
> that stuff should eat through anything nasty.
>
>
> Good luck,
>
>
> Mariena
>
> ________________________________
> From: 3dem <3dem-bounces at ncmir.ucsd.edu> on behalf of Lars-Anders Carlson
> <lars-anders.carlson at umu.se>
> Sent: Monday, July 1, 2019 1:23:10 PM
> To: 3dem at ncmir.ucsd.edu
> Subject: Re: [3dem] Help! Plastic films on my holey grids are KILLING ME
>
> Hi Cindi,
> Can you remove it by just plasma cleaning longer? That could be the
> reason why some of us typically don't see this film in the first place
> (another reason could of course be that we don't look as closely as you).
>
> QF grids used to have plastic deposits around the edge of the holes,
> which is where they would start bubbling at higher exposures (80-100
> e/?2), but I haven't seen much of this lately.
>
> I hope you find a solution since you seem really frustrated about it.
>
> Cheers,
> Lars
>
> On 2019-07-01 18:38, Schwartz, Cindi (NIH/NIAID) [E] wrote:
> > Hi all,
> >
> > We?ve been suffering with trying to get the plastic films off our
> > Quantifoil grids with very inconsistent results. We?ve also had film
> > issues on the c-flats, but my understanding is that the Quantifoils use
> > a plastic background and the c-flats use a carbon background. Please
> > correct me if I?m wrong on that one. We use both copper and gold grids
> > with the carbon substrate of various different patterns from R1/4 to
> > Multi-A with grids that we purchased 2 years ago to ones we?ve received
> > last month.
> >
> > The list of solvents we?ve tried that should dissolve plastic films:
> >
> > Ethyl acetate
> >
> > Chloroform
> >
> > Ethylene dichloride
> >
> > Trichlorobenzene
> >
> > The list of solvents to clean them up a bit:
> >
> > Acetone
> >
> > Ethanol
> >
> > 5% acetic acid
> >
> > We?ve tried sonication (a disaster)
> >
> > Submerging in drops
> >
> > Submerging in vast quantities of liquid
> >
> > With/without little stir bars while the grids are on a metal platform in
> > the solvent
> >
> > Time ranges from 15 min to overnight
> >
> > With/without a light glow discharge before cleaning
> >
> > We?ve put 10 nm colloidal gold fiducials on both sides of the grids
> > (like you?d use for plastic section tomography?where they stick to
> > Formvar just fine) after cleaning with various permutations from the
> > above lists and we see that the carbon has gold, and the ?holes? are
> > clean with no gold, only they aren?t clean! The colloidal gold does not
> > seem to stick to the plastic film because we see the film if we negative
> > stain apoferritin on these same grids or we see the films in the big
> > scope based on the ctf or the way the ice has blotted or how the films
> > have broken away and flipped onto the carbon. It?s rather obvious when
> > you also see true holes next to ?filmed? holes.
> >
> > Lastly, something will work for a subset of grids in a box, but later
> > on, grids from the same box (even the same SIDE of the box) don?t get
> > cleaned using the same protocol.
> >
> > All I can conclude is that whatever this plastic is, we should get
> > Quantifoil to stop using it because nothing seems to dissolve it and
> > it?s probably killing the planet somehow. If not, it?s certainly
> > detrimental to my mental health. Or better yet, Quantifoil should do the
> > research and either clean them better themselves or at least give us a
> > protocol to do so in the lab.
> >
> > Any tips, tricks, or surefire ways to clean grids would be great. And
> > please publish it because nothing pops up on Pubmed when I search for it.
> >
> > Thanks,
> >
> > Cindi L. Schwartz
> >
> > Electron Microscopist
> >
> > Rocky Mountain Labs/NIAID/NIH
> >
> > 903 South 4th Street
> >
> > Hamilton, MT  59840
> >
> > 406-363-9228
> >
> > Cindi.Schwartz at NIH.gov <mailto:Cindi.Schwartz at NIH.gov>
> >
> > Description: Macintosh
> > HD:Users:schwartzcl:Desktop:large-green-recycle-symbol-hi.png Please
> > remember our Earth before printing this email Description: Macintosh
> > HD:Users:schwartzcl:Desktop:large-green-recycle-symbol-hi.png
> >
> > *************************************************************************
> >
> > The information in this e-mail and any of its attachments is
> > confidential and may contain sensitive information. It should not be
> > used by anyone who is not the original intended recipient. If you have
> > received this e-mail in error please inform the sender and delete it
> > from your mailbox or any other storage devices. National Institute of
> > Allergy and Infectious Diseases shall not accept liability for any
> > statements made that are sender's own and not expressly made on behalf
> > of the NIAID by one of its representatives.
> >
> > *************************************************************************
> >
> >
> > _______________________________________________
> > 3dem mailing list
> > 3dem at ncmir.ucsd.edu
> > https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem
> >
>
> --
> Lars-Anders Carlson
> Assistant Professor
> Dept of Medical Biochemistry and Biophysics
> Wallenberg Centre for Molecular Medicine
> Ume? University
> 901 87 Ume?, Sweden
> http://www.carlsonlab.se/
> _______________________________________________
> 3dem mailing list
> 3dem at ncmir.ucsd.edu
> https://mail.ncmir.ucsd.edu/mailman/listinfo/3dem
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