Question

I'm trying to inspect simple stained bacterial smears. But my smear suddenly disappears after a successful inspection with the oil immersion lenses. The background can become too red (the color of the stain) and the oil residue coming from the oil immersion objective when it's wiped, is red colored. Also when I try to wipe the oil from the slide, the smear is removed, even with a gentle wipe. I'm suspecting three possible causes, which may include dependent causes: The immersion oil I use makes the smear easy to wipe:

I don't have a dropper that releases just a small drop, but I have used another oil with the same type of dropper that releases more than the needed quantity and it didn't wipe my smear.The back-and-forth movement that eliminates air bubbles: I hear the spring sound, is this correct?Smear fixation may not be adequate: could it be possible that the smear can be fixed in an incomplete way so that it resists being wiped from the staining procedure, but not from further manipulation? My specifications Immersion oil: Non-drying, non-hardening Cedarwood oilStain: Carbol fuchsin (20%)Smear fixation: heat fixation by passing the slide three times through the flame.

Answer 1

To achieve maximum resolution with the oil immersion objective, you must use a drop of oil. It is important to clean off the oil and lenses after using the microscope to prevent damage. Heat fixing the smear prepares it for staining with cationic or anionic dyes.

Step-by-step explanation:

It is important to remember that you must use a drop of oil whenever you use the oil immersion objective or you will not achieve maximum resolution with that lens. However, you should never use oil with any of the other objectives, and you should be diligent about wiping off the oil and cleaning all of your lenses each time you use your microscope, because the oil will damage the lenses and gum up other parts of the instrument if it is left in place.

The same thing happens as the light passes through the glass slide into the air space between the slide and the lens. The light will be refracted away from the lens aperture. To remedy this, we add a drop of oil to the slide and slip the oil immersion objective into it. Oil and glass have a similar refractive index, and therefore the light bends to a lesser degree and most of it enters the lens aperture to form the image.

Heat fixed smears are ready for staining. In a simple stain, dyes that are either attracted by charge (a cationic dye such as methylene blue or crystal violet) or repelled by charge (an anionic dye such as eosin or India ink) are added to the smear. Cationic dyes bind the bacterial cells which can be easily observed against the bright background. Anionic dyes are repelled by the cells, and therefore the cells are bright against the stained background.