Fungal Staining by Lactophenol Cotton Blue method (LPCB)

 Aim

To stain fungal cells by using Lactophenol cotton blue.

Principle 

The lactophenol cotton blue (LPCB) wet mount preparation is the most widely used method of staining and observing fungi and is simple to prepare. The preparation has three components:

1.      Phenol:   kills any live organisms;

2.      Lactic acid : It  preserves fungal structures, and

3.      Cotton blue : It stains the chitin in the fungal cell walls.

Lactophenol Cotton Blue Solution is a mounting medium and staining agent used in the preparation of slides for microscopic examination of fungi. Fungal elements are stained intensely blue.

Materials Required

1.      Microscope

2.      Glass Slide

3.      Cover Slip

4.      70 % alcohol

5.      Lactophenol cotton blue (LPCB)

6.      Inoculation needle

7.      Fungal culture

Procedure

1. Placed a drop of 70% ethanol on a clean microscopic glass slide
2. The Immersed the specimen in the drop of alcohol
3. Added  one or two drops of the LPCB before the alcohol dries out
4. Hold  the coverslip between the index finger and thumb, touch one edge of the drop of mountant with a coverslip edge and lower gently avoiding air bubbles
5. Made the initial examination using low power objective then Switched to higher power (40X) objective for more detailed examination of spores and other structures.

Observation

After staining fungus cell observed  different parts of a cell like conidia, hypae as well as conidiophore etc.

Result

On basis of observation the type fungal organism identified as Aspergillus sp.,