[3dem] Weird droplets

[bestdz at gmail.com]

Dear Sergej,
I believe those are (almost) dried out buffer of your sample. They are black because of high concentration of salt. You observe this because you have carbon film, otherwise they will be just empty holes. 
You should check your humidifier (less likely), or drastically reduce your blot time. If you are working with Vitrobot, and already have a very short blot time, say 1s, then you can try manual plunger, or concentrate your sample. Diluted samples are less viscous and thus easier to be blotted away. Also, the blot time should be much shorter when you have a carbon film. 
Good luck!

Best,

Xinghong Dai
Postdoctoral Scholar
Department of Molecular and Medical Pharmacology, UCLA
+1 310 447 4330

From: Sergej Masich
Sent: Wednesday, October 19, 2016 6:30 AM
To: 3dem at ncmir.ucsd.edu
Subject: [3dem] Weird droplets

Dear List, 

I probably missed the discussion since many of you had observed this phenomenon I suppose (see attached images). This is a Quantifoil grid with additional continuous layer of thin (5 nm) carbon. The grid was glow-discharged during 120 seconds, the solution was applied to the “proper” side (facing grid-box centre), blotted and plunge-frozen in liquid ethane. Why do these droplets form? I do not see the droplets after applying solution to the opposite side of the grid. Shouldn’t the carbon surface inside the holes be homogeneously hydrophilic? The surface outside the holes seems to be completely hydrophobic. Is it due to residual plastic?

Thank you in advance.

Sergej
=============================
Sergej Masich, Ph.D.
Dept. of Cell and Molecular Biology
Karolinska Institutet
171 77 Stockholm, Sweden
tel: (+)46 - 8 - 524 873 61
mobile: (+)46 - 736 - 833 693
e-mail: Sergej.Masich at ki.se




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