Smear Preparation basics

Inoculation and transfer techniques aseptically is covered in session 2.
Bacterial characterization includes differential media, biochemical tests and other tests in future sessions
Session 1 involves microscopy.
Session 3 invloves preparing slides for different staining treatments (simple, differential, special)

A loopful of water is added to the slide to create a suspension to spread the bactera
Add inoculum to the drop of water to create a suspension
Air dry the slide for less than 10 minutes
Heat fix the air-dried smear by passing thr slide through the flame about 3 times

Gram staining involves the use of crystal violet, iodine, alcohol, and safranin to stain the bacteria
Crystal violet stains all cells purple
Iodine acts as a mordant, helping to keep the primary stain bound to the cell
Gram-negative bacteria lose the primary stain upon rinsing with alcohol
Safranin is used as a counterstain
Gram-negative bacteria appears pink/red
Gram-postive bacteria appers purple

Used as a differential stain to distinguish organisms with waxy cell walls
Acid-fast bacteria contain mycolic acid, a waxy substance that makes it difficult for cells to react to gram staining
Mycolic acid makes the bacteria impermeable to some staining procedures
Cells that retain a basic stain when rinsed with acid-alcohol, are termed acid-fast positive cells
Cells will lose the stain when rinsed with acid-alcohol are termed non-acid-fast cells
Non-acid-fast cells are counterstained to see it in contrast with acid-fast cells

Uses carbol fuschin as the primary stain with phenol to help solubilize the cell wall
Heat is used as a mordant
Uses acid-alcohol to decolorize
Uses Methylene blue as the counterstain
Acid-fast cells appear red and non-acid-fast cells appear blue

Uses carbol fuchsin as the primary stain which has a higher concentration of phenol
No heat is sued
Uses acid-alcohol to decolorize
uses brilliant green as the counterstain
Acid-fast cells appear red, and non-acid-fast cells appear green

Prepare smears of organisms and heat fix the smear
Place slide on the staining rack then cover with a small paper towel soaked with carbol fuschin and allow to stand for 30 to 60 seconds
Heat gently until slight steaming is observed then maintain steaming for five minutes while adding more dye as needed to prevent drying of the smear
Cool teh slide then remove the paper and rinse with water
Decolorize slowly using an acid-alcohol then rinse with running water
Cover with methylene blue for one minute and rinse with water
Blot dry and observe

Etiologic agent of tuberculosis in humans and humans are the only reservoir for it
Leading cause of death in the world from a bacterial infectious disease
Affects 1.8 billion people/year which is a third of the world's population
More likely to be found in those with AIDS