Two -methods are generally used, (wet method) and (dry and fix method ) for Studying Microbes with a Compound Microscope
Wet Method: There are two primary methods generally used for studying microorganisms in wet conditions
wet mount method
hanging drop method.
The wet mount
A drop of fluid containing microorganisms.
The fluid spreads out in a thin layer between coverslip and slide.
The hanging Drop Method
It is used to observe the motility, germination, or fission of microorganisms.
Dry and fix method
Microorganisms, particularly bacteria, being too small need their permanent preparation be made by drying and fixing them on clean slide with or without staining.
For preparing a dry mount:
A drop of distilled water with a small amount of culture is spread as a thin smear on a clean slide. The smear is allowed to dry and it is then ‘fixed’ by passing it through a flame two to three times with the smeared slide away from the flame. If desired, this dried and fixed amount may be stained and the preparation dried again for observation under the microscope.
Bacterial Shapes :
Bacteria have three basic shapes:
Round cells (Cocci) , Rod-shaped cells are bacilli , spiral-shaped cells
Types of stain
Positive: basic stain
Attaches to cell wall and bacterium appears colored
Negative: acidic stain
Repelled from cell wall; stains background
To observe basic external structures of cell with brightfield scope (cellular morphology)
Reagent: Methylene Blue
Adherence to slide
Air dry first to prevent lysis (boiling)
Use of a single basic dye is called a simple stain.
A mordant may be used to hold the stain or coat the specimen to enlarge it.
The Gram stain classifies bacteria into gram-positive and gram-negative.
Gram-positive bacteria tend to be killed by penicillin and detergents.
Gram-negative bacteria are more resistant to antibiotics.
To view cellular morphology
Diagnostic purposes: What type of cell wall a bacterium has
Thicker peptidoglycan layer and no outer membrane
Thinner Peptidoglycan and an out membrane