Gram’s staining of curd/idly batter/yeast

Gram’s staining of curd/idly batter/yeast

Aim:

To determine the gram nature of microorganism present in the given sample(curd/ idly batter/yeast) by Gram’s staining technique.

Theory and Principle:

Based on gram staining reaction bacteria can be divided into two large groups called gram positive and gram negative. Gram positive bacteria have thicker peptidoglycan compared to the gram negative bacteria. The lipid content of the cell wall of gram negative bacteria is higher than that of gram positive bacteria. Both gram positive and gram negative bacteria take up the primary stain crystal violet and stain red. When decolorized the porosity and permeability of the gram negative bacteria increases due to which the crystal violet iodine complex is given out by the gram negative bacteria. Further the gram negative bacteria takes up the counterstain safranin and stains red. Hence when bacteria are observed under microscope after gram staining the gram positive cells appear violet and gram negative cells

Requirements:

• Clean grease free slide

• Nichrome loop

• Given culture

• Crystal violet

• Grams iodine

• Decolorizer(Acetone Alcohol)

• Safranin

• D/W

Procedure:

1. Take a loopful of the given culture and place on the slide.

2. Prepare a smear and heat fix it.

3. Cover the smear with Crystal Violet for one minute.

4. Wash gently .

5. Add Grams iodine for one minute.

6. Decolorise with acetone alcohol .

7. Wash the slide immediately.

8. Cover the smear with safranin for a minute .

9. Wash and Air dry.

10. Observe the slide under high power and oil immersion objectives.

11. Record your observations.

Diagram: (Any one Diagram)

Observation Table: ( any one shape and stain)

Results:

Gram staining of the given culture revealed gram positive violet colored rod-shaped bacteria in chains.