Simple staining

Aim

To study the morphological shapes, size and arrangement of bacterial cells

Principle

In simple staining the bacterial smear is stained with a single reagent.  The simple stain can be used as a quick and easy way to determine cell shape, size and arrangements of bacteria.   Most bacteria have a defined shape that falls into the following morphological categories. Spherical or cocci, straight rod or bacilli, spiral or spirochetes. A few bacteria are pleomorphic and can change their shape.  For example Arthrobacter. 

Simple staining is performed by using basic stains.  Basic stains with positively charged chromogen is preferred because bacterial nucleic acid and certain cell wall materials carrying negative charge strongly attracts and binds to the cationic chromogen.  The most commonly used basic stains are methylene blue, safranin and crystal violet. 

Materials required

1. 24 hour old bacterial culture - Bacillus and Staphylococcus species

2. Crystal violet/Safranin

3. Equipments - bunsen burner, inoculating needle, staining tray, microscope

Procedure

1.  The bacterial smears were prepared and heat fixed in a clean glass slide

2. The smear was flooded in crystal violet or safranin for 1 minute

3. Crystal violet/Safranin was washed off gently using slow running tap water

4. The smear was air dried and observed under oil immersion objective

Observation

Purple coloured cocci in cluster were observed on staining with Crystal violet. 

Red coloured rods were observed on staining with Safranin.

 

Result

The given sample contain purple coloured cocci in clusters and red coloured rods against colourless background