[3dem] How to make grids positively charged during or after glow discharge.

[Sacha De Carlo]

Oui :)

That’s the original but there more recent protocols maybe by Harris, with
step by step procedures.

Though the chemicals are poison and may not be used anymore; found on
researchgate:

"pentylamine or iso-amylamine" technique developed by Dubochet for the
protein-free spreading of DNA was substituted, because these amines are
poisons, with two others: by Robley Williams, 1977 ("poly lysine
technique", this is mentioned above) and by Keller (''BAC technique",
Vollenweider HJ, Sogo JM, Koller T. A routine method for protein-free
spreading of double- and single-stranded nucleic acid molecules. Proc Natl
Acad Sci U S A. 1975 Jan;72(1):83-7.). Both last methods work well and
reproducibly.
Good luck.

Sent from my mobile device
-----------------------------
Sacha De Carlo, Ph.D.
Business Development Manager EM
Dectris.com

On 3 Jul 2019, at 22:23, SCHMUTZ Marc (ICS) <
marc.schmutz at ics-cnrs.unistra.fr> wrote:


You mean that paper Sacha ?


A new preparation method for dark-field electron microscopy of
biomacromolecules1
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#aep-article-footnote-id1>
2
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#aep-article-footnote-id2>
3
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#aep-article-footnote-id3>
Author links open overlay panelJacquesDubochet4
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#!>
MichelDucommun4
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#!>Max
Zollinger
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#!>
EdouardKellenberger4
<https://www.sciencedirect.com/science/article/pii/S002253207180148X#!>

Journal of Ultrastructure Research
<https://www.sciencedirect.com/science/journal/00225320>
Volume 35, Issues 1–2
<https://www.sciencedirect.com/science/journal/00225320/35/1>, April 1971,
Pages 147-167

regards


Marc
------------------------------
*De: *"Sacha De Carlo" <sacha.decarlo at dectris.com>
*À: *"Ross, Bradford" <bradford.ross at botany.ubc.ca>
*Cc: *3dem at ncmir.ucsd.edu
*Envoyé: *Mercredi 3 Juillet 2019 22:09:02
*Objet: *Re: [3dem] How to make grids positively charged during or
after        glow discharge.

You would need to read the old classics by people trying to look at dna in
negative staining: look for old protocols using pentylamine (if I find
them, I will share them with you) but I’m sure there are some review papers
too.
Best
Sacha


Sent from my mobile device
-----------------------------
Sacha De Carlo, Ph.D.
Business Development Manager EM
Dectris.com

On 3 Jul 2019, at 20:06, Ross, Bradford <bradford.ross at botany.ubc.ca> wrote:

Hello Everyone,


I'm currently trying to image some lipid nanoparticles which have a
positive charge on their outer surface. Of course, since normal glow
discharge treatment typically yields a negative charge on the grids, the
particles are all sticking to the lacey carbon support instead of remaining
suspended in the vitreous ice layer.


I have heard that Magnesium Acetate can be used to switch the charge from
negative to positive after glow discharging, but can't seem to find any
protocols for exactly how to treat the grids with it.


Does anyone have a protocol for the use of Magnesium Acetate, or any other
suggestions on how to flip the surface charge of the grids to positive? Or
other creative solutions I'm not thinking of?


Cheers,
Bradford Ross

Electron Microscopy Technician
BioImaging Facility
University of British Columbia
Cunningham Building Rm. 64
2146 East Mall
Vancouver, B.C.
V6T 1Z4

phone 604-822-6996

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