WANT TO SEE A BACTERIUM UNDER THE MICROSCOPE? USE THE GRAM STAIN

Well it is easy to use the different stains like H&E , Nile blue sulphate, or Neutral red or Leishman to stain different cells and observe them. But does it apply the same for bacteria? l forgot to mention that bacteria are the largest numbers of life forms on this planet, existing form within your intestine to the deep underwater vents of hot gases under the ocean or the Tundra! It is also interesting to note that all bacteria can be basically divided into two groups on basis of their staining properties by a stain called Gram stain, discovered by  Hans Christian Gram in 1882–1884.

  • Gram stain is based on the physical and chemical differences of cell wall in bacteria
  • Bacteria  can be categorized into two types-Gram positive  and Gram negative.

SAINING PROCEDURE:

  • Heat fixed sections are placed in distilled water
  • The  section is flooded  with Crystal violet or the Primary stain, as it colorizes all types of cells .
  • Crystal violet is washed off after 1 minute with distilled water.
  • The section is  flooded Grams iodine solution , which is called a mordant or a helper.
  • Iodine is washed off  after approximately a minute.
  • The slide is washed with 95% alcohol or alcohol -acetone solution  to remove excess purple from some bacteria, it is decolorization agent.
  •  The slide must be kept at an angle and  alcohol   has to be poured till there is no purple tint.
  • Slide is washed immediately with tap water.
  • Slide is counter stained with safranin for 1 min.
  • It is washed with  distilled water, air dried and mounted for observation.

OBSERVATIONS:

  1. Blue  or Black-        Indicates gram positive organisms
  2. Blue-                           Indicates fungal filaments
  3. Red-                            Indicates gram negative microorganism
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