megha sharma wrote:
> I am working with gram negative magnetotactic bacteria (micro aerobic) and i
> wish to stain then fluorescently. The contain bilipidlayer
> organelles (magnetosomes) which may be empty or cantain nano magnetite
> particles.
> I need the stain to improve the contrast while making videos of the motile
> cells
>> I was also wondering how can I count the motile cells under the microscope
I haven't done a lot of work with microscopy, so I won't be able to
offer a lot of help. But, here's what I do know:
If you are interested in keeping your cells alive, then you have to be
careful about which staining procedure you pick. A lot of them (such
as Gram staining) kills the cells. However, I think you can do
fluorescent antibody staining without killing the cells. You would
need to order antibodies that are specific for some outer structure on
your bacteria, and the antibody also has to have some type of
fluorescent dye attached to it. That's one way.
Another way would be to transform a plasmid expressing GFP (green
fluorescent protein) into your cells. This would cause them to glow
green. I believe this would make videos and counting quite
nice....especially with a dark background (darkfield microscopy, I
believe).
Anyway, as I said, I do very little microscopy work. I'm sure other
people on here know a lot more about this than I do.
--Alex
************************
Alex B. Berezow, Grad Student
Dept. of Microbiology
University of Washington School of Medicine
Seattle, WA 98195
