Basic stain
Cationic-chromogen exhibits poss charge
Strong affinity to neg charge cell components
More commonly used for bacterial satining.
Acidic stain
Anionic-chromogen portion has neg. charge
Strong affinity for positive components of cell
Acid fast
Stain used for pathogenic bacteria for humans such as M.tuberculosis and M.leprae.
Acid fast stain
Primary stain- carbol Fuchsim
Decolorizing- acid alcohol
Counter stain- methylene blue
Color of acid fast
P-Carbol Fuchsim- red
D-Acid-alcohol- red
C-Methylene blue- red
Color of Non acid fast
P-Carbol Fuchsim-Red
D-acid alcohol- colorless
C-Methylene blue-blue
Endospore stain
Primary stain-Malachite green
Decolorizing agent- water
Counter stain- safranin
Endospore Primary stain
Malachite green- flood smear for 2-3 minutes over warm hot plate
Decolorizing agent
Water- removes excess malachite green.
once spore accepts green, it can’t be decolorized by water.
Counterstain
Safranin-red stain used as second reagent.
Colors decolorized vegetative cells, appear read after absorption of counter stain
Spore stain- color of stain
P-malachite green-Green
D-water- Green
C-Safranin-Green
Spore stain-Color of vegetative cell
P-Malachite green-Green
D-Water-Colorless
C-Safranin- Red
Spore staining procedure
1) flood smear with malachite and place on top of warm water beaker for 2-3 min. Don’t let it evaporate
2) remove slide from beaker, cool, and wash with tap water.
3) Counterstain with safranin for 30 sec.
4) Wash with tap water
5) dry slide with bibulous paper.
Gram stain
Primary stain-crystal violet
Mordant-Iodine
Decolorizing agent- alcohol-acetone
Counter stain-safranin
Gram Stain procedure
1) gently flood smear with crystal violet for 1 min
2) gently wash with tap water
3) gently flood smear with Iodine mordant for 1 min.
4) gently wash-tap water
5) decolorized with alcohol-acetone
6) wash with tw
7) Counterstain safranin for 45 sec.
8) wash with tw
9) dry with bibulous paper and examine oil immersion
Color of G+ cells
P-Crystal violet- purple
M-Iodine- purple
D- alcohol-acetone- purple
C- Safranin- purple
Color of G- cells
P-crystal violet- purple
M-iodine- purple
D-alcohol acetone- colorless
C-Safranin- red
Simple staining
Single reagent smear. Produces distinct contrast between organism an it’s background.
Simple staining procedure
1) flood smear with indicated stain Carbol Fuchsim 15-30 sec Crystal violet 20-60 sec Methalyne blue- 1-2 min 2) wash smear with tw 3) dry using bibulous paper 4) repeat procedure for 2 rem. stains 5) oil immersion