Lab 5: Differential Staining (Gram Stain and Acid-Fast Stain)

Flashcards covering key vocabulary terms related to Gram staining and acid-fast staining procedures, including reagents, cell wall characteristics, and examples of bacteria discussed in Lab 5.

Differential Staining

Staining procedures that differentiate or separate cells based on staining differences, allowing detection of structural variations or identification of parts of an organism.

Gram Stain

One of the most widely employed staining techniques in microbiology that separates bacterial cells into two types (gram-positive and gram-negative) based on structural differences in their cell walls.

Gram-Positive Cells

Bacterial cells characterized by a thick peptidoglycan cell wall, retaining the crystal violet-iodine complex and appearing violet-purple after Gram staining.

Gram-Negative Cells

Bacterial cells characterized by a thin peptidoglycan cell wall and an outer membrane, losing the primary stain during decolorization and appearing red-pink after counterstaining with safranin.

Crystal Violet

The primary stain in the Gram stain procedure, a basic dye that initially stains both gram-positive and gram-negative cells purple.

Gram's Iodine

The mordant used in the Gram stain procedure, which binds to crystal violet forming an insoluble complex trapped within the cell.

Decolorizer (Gram Stain)

A mixture of alcohol and/or acetone (typically 50:50 ethanol and acetone) used in Gram staining to remove the primary stain from gram-negative cells.

Safranin

The counterstain in the Gram stain procedure, a red-pink dye that stains gram-negative cells after they have lost the primary stain, causing them to appear red-pink.

False Gram-Negative Result

An error in Gram staining where gram-positive cells incorrectly appear gram-negative, often caused by using cultures older than 24 hours (due to weakened cell walls) or over-decolorization.

False Gram-Positive Result

An error in Gram staining where gram-negative cells incorrectly appear gram-positive, often caused by a smear that is too thick, trapping crystal violet.

Acid-Fast Stain

A differential stain technique used to detect the presence of waxy lipids called mycolic acids within a bacterial cell wall.

Mycolic Acid

Waxy lipids found embedded in the thick cell walls of certain bacteria (e.g., Mycobacterium and Nocardia), making them resistant to general staining and harsh environments.

Acid-Fast Cells

Bacterial cells containing mycolic acid that retain the primary stain (red carbolfuchsin) even after decolorization with acid-alcohol, appearing red.

Non-Acid-Fast Cells

Bacterial cells lacking mycolic acid which lose the primary stain after decolorization and are subsequently stained blue by the counterstain (methylene blue), appearing blue.

Carbolfuchsin

The primary stain in the acid-fast staining procedure, a dark red, lipid-soluble stain that penetrates waxy mycolic acid in cell walls, often aided by heat.

Heat (Acid-Fast Stain)

Applied during the primary staining step of the acid-fast procedure to melt the mycolic acid, facilitating the penetration of carbolfuchsin into the cell.

Acid Alcohol

The decolorizer used in the acid-fast staining protocol, a combination of ethyl alcohol and hydrochloric acid, which removes the primary stain from non-acid-fast cells.

Methylene Blue

The counterstain used in the acid-fast procedure, which stains non-acid-fast cells blue after they have been decolorized.

Mycobacterium

A genus of bacteria known to contain mycolic acids in their cell walls, making them acid-fast (e.g., M. tuberculosis, M. leprae, M. smegmatis).

Nocardia

A genus of bacteria that, like Mycobacterium, possesses mycolic acids in its cell walls and is identified using the acid-fast stain.

Cording (Mycobacterium)

A characteristic arrangement where mycolic acid causes bacterial cells to stick together, forming large masses or 'cords'.

Bacillus subtilis

A large gram-positive bacillus (rod-shaped) that typically forms chains (streptobacillus) and is found in soil, usually non-pathogenic.

Mycobacterium smegmatis

A weak gram-positive bacillus that is also acid-fast due to the presence of mycolic acid; it forms cords and is non-pathogenic, found in soil, plants, and water.

Staphylococcus aureus

A gram-positive coccus that characteristically forms clusters (staphylo arrangement) and is a known human pathogen capable of causing various infections; BSL-2 organism.

Streptococcus salivarius

A gram-positive coccus that typically forms chains (strepto arrangement); it is often found in the oral cavity.

Escherichia coli

A gram-negative bacillus (rod-shaped) that typically resides in the gastrointestinal tracts of animals; while some strains are pathogenic (e.g., O157:H7), lab strains are often BSL-1.