Spore stainingIn addition to malachite green staining, are there other staining methods?

It can also be used with carbolic acid to re-red stain.

The principle is that the spores have a thick wall and low permeability and are not easy to color, but once the spores are colored, it is difficult to be decolorized. Therefore, a dyeing solution with strong coloring ability (such as malachite green or carbolic acid re-red) is first used to stain under heating conditions, so that the dye can enter both the cell and the spores, and the dye in the cell is eluted when the water is eluted and decolorized, while the dye in the spores is still retained. After staining with a high-contrast counterstain, the bacteria are dyed with the color of the counterstain, and the spores are still the original color, so that the two are distinguished.

I don't know, I've never heard of this method

1) Take two clean small test tubes, add 0 2ml of sterile water respectively, and then add 1-2 inoculation loops of Bacillus cereus to one tube, and add 1-2 inoculation loops of Clostridium sporigenes to the other tube, and the two tubes are fully mixed into a thick bacterial suspension. (2) Add 0 2ml of phenol fuchsin solution to the bacterial suspension, mix it thoroughly, and heat it in a boiling water bath for 3-5 minutes. (3) Take 2-3 rings of the above mixture on two glass slides with inoculation loops, coat them thinly, air dry, tilt the slides slightly on the beaker, and rinse them with 95 ethanol until no red liquid flows out.

4) Rinse with tap water and blot the filter paper. (5) Take 1-2 inoculation loop melanin solution on the smear, immediately apply thin, after natural drying, observe with oil microscope, under the background of light purple gray, the bacteria are white, and the smolders in the bacteria are red.

1. Why do I need to heat for dyeing?

The most stress-resistant life organism in the whole biological world, whether it can destroy the spores is the most important indicator to measure the various disinfection and sterilization methods, only in the process of heating the permeability of the spore has changed, the dye liquid can penetrate into the spores, so the spore staining needs to be heated.

2. If only free spoblasts are seen in the production, but few spore sacs and vegetative cells are seen, why?

If only spores are seen, it could be:

1）.The culture environment is too harsh, and the vegetative bodies are almost all about to turn into spores.

2）.It may be that the spore staining was not washed with water, and the cells are still malachite green.

3）.If there is a problem with the re-ignition, it may be that the re-ignition time is not enough, it is not dyed, or the water is too badly washed and the dye is washed away.

Hope it can help you,

Because the bacteria has been cultured for too long, it has reached the middle and late stages of growth. With the depletion of nutrients in the environment and the accumulation of secondary metabolites in bacteria, the environment is becoming less and less suitable for bacteria to grow, so spores must be produced to survive the adverse environment. As a result, a large number of spores are produced, and at the same time, the sporangia disintegrate.

The simple staining method is suitable for the observation of the general shape of the cell and the arrangement of bacteria. Simple dyeing process:

1) Smear: Take a clean glass slide and drop a small drop of normal saline (or sterile distilled water) in it. After sterilization on an alcohol lamp, the inoculation loop picks a small amount of bacteria (Staphylococcus aureus or Bacillus subtilis) from the inclined plane and mixes with the water droplets on the slide.

No. Because the simple dyeing method can not color the spores, the spore wall is thick, the permeability is low, and the coloring and decolorization are more difficult, and the spore staining is generally based on the principle that the spores of the bacteria and the bacteria have different affinities for the dyes, and different dyes are used for coloring, so that the spores and the bacteria are different colors and easy to distinguish.

(1) Select the strain of appropriate age, the young bacteria have not yet formed spores, and the sporangia of the old bacteria have been ruptured.

2) Heating and dyeing must be maintained in the state of steam in the dyeing solution, heating and boiling will lead to the rupture of the bacterium or sporangium, and it is difficult for the spores to be colored if the heating is not enough.

Introduction: Methods of spore staining:

1) Bacillus subtilis or other bacillus bacteria that have been cultured for about 24 hours are smeared, dried and fixed.

2) Add 3-5 drops of malachite green stain solution to the fixed smear.

3) Hold the glass slide with a wooden clamp and heat it on the flame to make the dye steam but not boil, do not evaporate the dye dry, and add a little dye if necessary. The heating time is calculated from the time when the dye is steaming, about 4-5 minutes. This step can also be done without heating and used saturated aqueous malachite green solution (about 10 minutes.

4) Pour off the staining solution, wait for the slide to cool and wash with water until the malachite green no longer fades.

5) Counterstain with angustron aqueous solution for 1 minute, and wash with water until the water is colorless.

6) After drying, the spores are green and the bacteria are red.

Spore is a highly stress-resistant hypnozoite structure formed by Bacillus and Clostridium spp. at a certain stage of growth, usually round or oval in shape. The spore wall is thick and permeable, and it is difficult to colorize and decolorize. Therefore, when dyeing with a dyeing agent with strong dyeing power (such as malachite green) under heating conditions, the dye can not only enter the bacterium, but also enter the spores, the dye that enters the bacterium can be eluted and decolorized by water, and the dye that enters the spores is difficult to penetrate, if the counterstaining solution (such as sand yellow aqueous solution) is stained, the spores still retain the color of the primary dye, and the bacteria are dyed into the color of the counterstain, that is, the bacteria and spores are dyed red and green respectively and are easy to distinguish.

The chemical reagent used to perform spore brigade first staining is () tie sensitization.

a.Neutral red.

b.Crystal purple knowledge town branches.

c.Cotton blue. d.Malachite green.

Correct answer: d