I am using a conformational dye that bonds to some structures on bacterial membranes and formaldehyde fixation does not really work for me.
Thanks!
There are several publications in the Journal of Bacteriology in which cryo fixation of bacteria was used.
Good Morning,
I recommend that you review the following literature.
Chapter 2 – Bacterial TEM: New Insights from Cryo-Microscopy
Martin Pilhofer*, †,
Mark S. Ladinsky*,
Alasdair W. McDowall*,
Grant J. Jensen*, †
* Division of Biology, California Institute of Technology, Pasadena, California 91125
† Division of Biology, Howard Hughes Medical Institute, Pasadena, California 91125
hi
Use 4% paraformaldehyde (PFA) to fix all sorts of cells, for cells in culture incubation for 15-20 mins is sufficient to fix. You will need to heat it up to dissolve PFA in 0.1M PBS and adjust pH to 7.4. Needs to be done under the fumehood. Need to use it fresh, but OK for a few days in the fridge, or longer in freezer (make sure you use a seperate (dedicated) fridge/freezer as PFA can release fumes that may effect other solutions/cells.
Once you fix your cells at room temperature for 15-20 mins than you need to wash the fixative off with 0.1M PBS.
Fixation basically stabilizes all the proteins including the one in the membrane and therefore it will not change the integrity of membranes, and Rifampicin should stay in the cell.
Hope this helps
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Cell Biology