Practical Basic Microbiology and Immunology. Microbiology. Microbiology is the science dealing with microorganisms. We are going to study microorganisms: Microscopically Macroscopically. Our session today. Culture media Aseptic technique Sources of contamination
-A source of energy.
-Sources of carbon, nitrogen, sulfur, phosphorus.
-Minerals, e.g., Ca2+ ,Mg2+,Na+.
-Vitamins and growth factors.
Nutrient broth :
- Composed of nutrients + water.
-Similar to nutrient broth but supplemented with solidifying agent (1-2% agar).
Plate (Petri dish)
How to avoid contamination of our work with pure culture?
1- Working in 20 cm diameter around a blue flame (sterile zone).
2- Never leave a culture dish open, even for a short time ,when it is necessary to open a dish, keep the lid close to the dish, and keep the lid between your face and the agar surface.
3- For most bacterial cultures you will use a sterile loop or needle to inoculate or to obtain an inoculum.
4- Flame a loop or needle to red-hot just prior to use, burning off any organic material, cool the loop prior to touching a culture.
one sterile Petri-dish
one molten nutrient agar tube
1- Pour the nutrient agar tube at the suitable temperature aseptically into the provided plate.
2- Leave the nutrient agar plate to solidify.
3- Expose the agar plate to one of the following source of contamination:
- Air (outside the aseptic zone)
- Forced air
- Skin touch
- Cough (Inside the aseptic zone)
- A piece of hair or cloth
- Drop of water
4- A student in each bench will perform a control plate.
5- Label the cover of the plate with your name, no., source of contamination.
6- Incubate the plate inverted , except for water containing plates.
1- A circle should be marked on the under side of a slide with a permanent marker.
2- To prepare a smear from a suspended culture, by means of an inoculating loop, aseptically transfer 3- 4 loopfuls of the culture (after shaking), place directly on the slide and spread gently in 1 cm area.
3- Air dry or dry over the flame.
Dried bacterial smear
4- Heat fixation : By passing the slide an inoculating loop, aseptically transfer 3- 4 loopfuls of the culture (after shaking), place directly on the slide and spread gently in 1 cm area.
about five passes through the flame.
5 to 6 times
Fixation while Incomplete drying Or Too much fixation
Too little fixation
Scheme for description of M.O