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Gram Staining

Gram staining

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shivamhero65
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11 views

Gram Staining

Gram staining

Uploaded by

shivamhero65
Copyright
© © All Rights Reserved
Available Formats
Download as PDF, TXT or read online on Scribd
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Gram staining is a widely used and trusted technique to identify bacteria on the

basis of cell wall composition. It is a differential staining technique used to


differentiate between Gram positive and Gram negative bacteria on the basis of cell
wall structure. Gram staining involves a series of steps, which are briefly
described below:

1. A smear of bacterial culture is prepared on a slide.

2. The smear is then fixed on the slide by heat or a chemical fixative such as
methanol or ethanol.

3. The smear is then stained with a purple dye called Crystal Violet.

4. After staining, the slide is washed with water to remove the excess dye.

5. Then the slide is treated with iodine solution, which acts as a mordant and
helps to fix the dye to the bacterial cell wall.

6. The slide is then washed with alcohol or acetone, which decolorizes the Gram-
positive bacteria and leaves the Gram-negative bacteria purple.

7. Finally, the slide is counter-stained with a pink or red dye such as safranin,
to give contrast between the Gram-positive and Gram-negative bacteria.

Gram staining works by exploiting the differences in the cell walls of Gram-
positive and Gram-negative bacteria. Gram-positive bacteria have thick and rigid
cell walls composed of a single layer of peptidoglycan, which is able to retain the
crystal violet dye. On the other hand, Gram-negative bacteria have much thinner and
more porous cell walls, composed of a thin layer of peptidoglycan surrounded by an
outer membrane which does not retain the crystal violet dye. When the slide is
decolorized with alcohol or acetone, the thick cell wall of Gram-positive bacteria
retains the crystal violet dye, while the thin cell wall of Gram-negative bacteria
is unable to retain the dye and is decolorized. The slide is then counter-stained
by a pink or red dye, such as safranin, to provide contrast between the Gram-
positive and Gram-negative bacteria.

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