Differential Stain- Gram Staining Flashcards
Purpose gram staining technique
to distinguish bacteria into Gram Positive or Negative based of the structure of the bacterial cell walls
Gram Staining Technique
Primary Stain (Crystal Violet) , Mordant (Gram’s Iodine), Decolorizing Agent (Alcohol), Counterstain/ Secondary Stain (Safranin)
Gram Staining Exact Techniques w/ Times
CV for 30 SECONDS, rinse w/ distilled water, Grams’s Iodine 10 SECONDS, rinse w/ distilled water, Ethanol for 12 SECONDS, rinse with distilled water, Safranin for 30 SECONDS, rinse w/ distilled water. Blot it dry
The first step/technique used to identify bacteria is
Gram Staining
Gram Positive cells stain
Purple
Gram Negative Cells stain
Pinkish/ Reddish
The primary stain (Crystal Violet) in gram staining, stain inside of cells because
Crystal Violent is a basic dye
How does the decolorizing agent (alcohol) affect Gram Negative Cell?
The decolorizing agent damages the outer membrane (LPS layer) of the Gram Negative Bacteria. Its thin layer of peptidoglycan cannot hold back the CV- Iodine dye complex.
Why do Gram Positive cells remain purple?
Their cell wall prevents the crystal violet - iodine complex to be washed out during decolorization.
How does the dehydrating agent (Alcohol) effect the cell wall of Gram positive cells?
The agent dehydrates the thick layer of peptidoglycan, and the cell wall acts as a permeable barrier that prevents the dye from leaving the cell
Bacteria that decolorize easily and become stained red
Gram Negative
Bacteria that decolorize slowly and retain primary stain (CV that stains purple)
Gram Positive
The mordant, Gram’s Iodine does what with primary stain Crystal Violet?
It forms a complex that is less likely to be washed away from the cell
Before any types of dyes, cells are typically
colorless
Before any types of dyes, cells are typically
colorless
