NURSING ANM AND GNM

Detailed explanation-1: -The alcohol will decolorize the sample if it is Gram negative, removing the crystal violet . However, if the alcohol remains on the sample for too long, it may also decolorize Gram positive cells. Add the secondary stain, safranin, to the slide and incubate for 1 minute.

Detailed explanation-2: -Gram-negative bacteria stained with crystal violet are decolorized by 95% alcohol within 2 min, whereas Gram-positive bacteria require at least 3 min treatment.

Detailed explanation-3: -Gram-negative cell walls contain a high concentration of lipids which are soluble in alcohol. The decolorizer dissolves the lipids, increasing cell-wall permeability and allowing the crystal violet-iodine complex to flow out of the cell. The color of the counterstain must contrast with that of the primary stain.

Detailed explanation-4: -After decolorization, the gram-positive cell remains purple in color, whereas the gram-negative cell loses the purple color and is only revealed when the counterstain, the positively charged dye safranin, is added.

Detailed explanation-5: -The organisms that do not take up primary stain appear red under a microscope and are Gram-negative organisms. The final step in gram staining is to use basic fuchsin stain to give decolorized gram-negative bacteria pink color for easier identification. It is also known as counterstain.