Gram stain troubleshooting

[llbutters]

Hi I've been trying to perform a gram stain of some liquid bacterial cultures but when I look at the slides under a microscope there is nothing there. It seems to be the iodine that's washing away the smear but I can't be sure. Can anyone help?

[Micro.Pete]

Hi there,

My guess is that your fixation wasn't perfect. Smear your bacteria on a microscope slide and wait until it is dry (you can put your slide in an incubator to speed up the drying process). After that you have to fix the bacteria to the slide. The easiest way to do it is to place the bottom side of the slide under a flame, such as a lighter or a bunsen burner. Just Make sure you don't burn your bacteria (the whole process should be quick).

 

Hope that I helped.

[MonDie]

I'm probably way way way underqualified for this, but could they be acid-fast? Acid-fast stain.

 

The high mycolic acid content of certain Protozoa cell walls, and those of Mycobacteria, is responsible for the staining pattern of poor absorption followed by high retention.

 

http://en.wikipedia.org/wiki/Acid-fast

 

Recently, two groups have reported that specimens containing many mycobacteria resulted in a remarkable gram-neutral (neither positive nor negative) or gram ghost appearance of the mycobacteria;

 

 

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC269558/?page=1

 

 

Endospores won't stain either, but I don't know how you would test for them.

Edited March 12, 2015 by MonDie