Why are Gram positive and Gram negative bacteria always seen under oil immersion microscope?

Answer 1

microman is correct.

I am a microbiologist. We can often see (at least some) bacteria, without using oil immersion examination, but you cannot see precise details of their shape and arrangement, and these details are important clues about what types they might be.

Small Gram Negative bacteria, such as Haemophilus influenzae and Bacteroides fragilis, and helical Gram Negatives such as Campylobacter species, might not be noticed, without the better magnification allowed by oil immersion microscopy.

Hey!! I sometimes have to look for several minutes to see these organisms, in clinical samples; the difference in colour and refraction, between the bacteria and the mucus etc of the sample, can be quite subtle, sometimes. I even failed to see a Fusobacterium in a sample of pus, a year or so ago; I said "No organisms seen", on the direct microscopy, and the bug grew all over the place! I looked again, using my "retrospectoscope", and (hindsight being a marvellous thing!) there they were; all over the sample ... just so very pale and thin, that I had missed them. They were still only really visible at the very edge of the smear, BTW.

:-)

Answer 2

You can still see both these types of stained bacteria even without using oil immersion. The reason immersion is used is because it increases the refractive index of the light entering a high-powered microscope lens. Lenses can only have a numerical aperture of 1.0 when it is separated from the slide by air. The aperture however increases with oil.

Answer 3

In Gram-positive bacteria, the purple crystal violet stain is trapped by the layer of peptidoglycan which forms the outer layer of the cell.
Hence, it is going to be purple.


In Gram-negative bacteria, the outer membrane prevents the stain from reaching the peptidoglycan layer in the periplasm. The outer membrane is then permeabilized by acetone treatment, and the pink safranin counterstain is trapped by the peptidoglycan layer.

So, it will show up as pink.
So it is seen under microscopes (general and oil immersion)

Answer 4

not always but *best* seen ... because they are very small and oil immersion gives the highest magnification with light microscopes ...

Answer 5

lysozyme ia a protease which may cut the derived proteins, so NOT suitable. detergent or surfactant also cause structural disturbances and denaturation of proteins so not suitable. anitbiotics, besides being expensive are hard to separate from the broth in later stages, so not suitable. transpeptidase in fact cross links and aids in forming cell walls so cannot be used for separating the two groups. EDTA is therefore the most preferred method for separating the two groups on lab scale.