PRE-ANALYTICAL PHASE

Sterilization of working area is very necessary

by using Lysol or bleach to wipe the tabletops.

GRAM STAINING
The laboratory scientist should always ware
PERSONAL PROTECTIVE EQUIPMENTS at all
times inside the laboratory and considered all
TECHNIQUE specimens infectious.

ANALYTICAL PHASE PROCEDURE

Materials 1. Prepare bacterial smears from the bacterial suspensions given. 2. Flood
the smear crystal violet for one minute.
Broth cultures 3. Wash off excess stain gently with running water.
Slides 4. Flood the smear with Gram's Iodine for one minute.
5. Wash off the Gram's Iodine with running water.
Inoculating 6. Decolorize the smear by flooding with 95% alcohol. Allow it to stand for
loops 15-30 seconds. Repeat this procedure until no more color comes off with
Crystal Violet the alcohol.
7. Wash again with running water.
Gram's iodine 8. Counter-stain with Safranin for 30 seconds
95% alcohol 9. Wash off the excess stain with running water. Air dry.
10. Examine the stained smear under oil immersion objective (OIO).
Safranin
GRAM (+) becomes (-)
Over-decolorization
Old dying
Acidic iodine

GRAM (-) becomes (+)

Under-decolorization
Thick smear
POST-ANALYTICAL
After the experiment, all the glassware, materials that have been
contact with bacteria like inoculating wire and including the
working station, should be sterilized properly. The student must
prepare 10% bleach or Lysol for disinfection. Materials like
inoculating wire can also be sterilized by famine, all disposable
END
materials should be place properly in correct trash bags.

PPE should be disposed in infectious trash bag, and should not be

worn outside the laboratory