When preparing a slide for a negative stain Why would you heat fix?
When preparing a slide for a negative stain Why would you heat fix?
While the slides are drying, start your negative stain. 6. Once the liquid has completely evaporated, heat fix by passing the slide through a flame three times. If you heat fix too little, the bacteria will wash off the slide.
Why did you not heat fix this slide?
Most bacteria have some kind of CAPSULE. Most bacterial capsules are composed of polysaccharide however some genera produce polypeptide capsules. Capsular material is very moist (slimy) and any heating will cause it to shrink – it is for this reason that we will not heat fix the slide before staining.
Why is it necessary to heat the slide after the smear is air dried?
Air-drying adheres the bacteria to the slide. Air-drying ensures that the smear is thin enough to stain. The air-drying step coagulates the proteins in the bacteria. During heat fixation, it is important that the slide be passed only 2-3 times through the outer portion of the Bunsen burner flame to prevent overheating.
Why is it important to let the slide dry completely before it is heat-fixed?
If you don’t completely air dry your slide, you will end up boiling your microorganisms when heat-fixing. When heat-fixing, if you hold the slide over the flame too long, you will incinerate the cells.
What happens if you forget to heat fix your slide?
You forgot to heat-fix your slide. Heat-fixing the slide fixes the bacteria to the slide surface. If this step is not done, the bacteria in the smear would be washed off of the slide during the staining and decolorization steps. You would not see anything on the slide under the microscope.
What would be some of the consequences for not leaving the stain on the slide long enough?
What are some consequences of not leaving a stain a smear long enough (under-staining)? Some consequences of under-staining are that the cells may lose their stain when washed with water or alcohol. This would make it difficult to identify the cell under the microscope.
What are the two purposes of heat fixation?
Two purposes of heat fixation are: Kill the microbes. Living bacteria contain enzymes that can breakdown the structures of the bacteria.
What is heat fixation in microbiology?
Heat fixation: After a smear has dried at room temperature, the slide is gripped by tongs or a clothespin and passed through the flame of a Bunsen burner several times to heat-kill and adhere the organism to the slide. Heat fixation generally preserves overall morphology but not internal structures.
What are three main reasons for heat fixing an organism to a slide?
Discuss the reasons for heat fixing bacterial smears prior to staining.
- To kill bacteria on slide;
- to make dye penetrate cells;
- to coagulate proteins of cells thus making cells stick to slide.
Does heat fixing distort cells?
Heat fixation tends to damage and distort the delicate cell wall structures of bacteria. This distortion also happens with tissue and blood cells which can create background debris that may be confusing or misleading.
What is the mechanism of negative stain?
Principle of Negative Staining Negative staining requires an acidic dye such as India Ink or Nigrosin. India Ink or Nigrosin is an acidic stain. This means that the stain readily gives up a hydrogen ion (proton) and the chromophore of the dye becomes negatively charged.