how to attach particles to carbon film on grid for EM imaging? We tried poly lysine,but it will contribute contrast for EM imaging
Dear Xusan,
In the past, when I did negative staining of DNA on grids, I used poly-lysine. Since I do not make DNA origami, I did a quick PubMed search which suggests that people just adsorb their sample onto a carbon-coated grid without poly-lysine. However, they make sure that the grids are freshly negatively glow-discharged.
The most complete materials and methods are in the supplemental materials for these two papers:
http://www.ncbi.nlm.nih.gov/pubmed/21493857
http://www.ncbi.nlm.nih.gov/pubmed/26149412
Good Luck!
Jill
We use carbon film TEM grid that are freshly negatively glow discharged and deposit the DNA origami sample solution directly on it. I guess the glow discharge cleans the grid surface and give the carbon film a slight negative charge (somewhat hydrophilic) so that the buffer can spread out and DNA origami can attach to the surface possibly mediated though the Mg2+ ion present in the buffer.
Many thanks for Jill Miotkeand Yan Liu's reply. glow discharge really can make origami attach to the surface well,however,the fluo molecule will be quenced bycarbon film？ We should use different kind of grid with different coating？
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