Bacterial Staining Practice Flashcards

A comprehensive set of practice questions covering bacterial staining principles, smear preparation techniques, and specific staining methods like Gram, Acid-Fast, and Spore stains.

What are the three steps in the bacterial staining workflow?

Smear preparation -> Staining -> Microscopic Examination/Visualization

What is the charge of the chromogen in an Acid Stain (dye)?

Anionic (negativity)

How do stains generally bind to bacterial cells?

Through electrostatic attraction, where opposite electrical charges attract (Positive $\leftrightarrow$ Negative).

What is the mnemonic BPAN used for in staining?

B = Basic, P = Positive, A = Attracts, N = Negative (Basic stains are positive and attract negative bacterial cells).

What are the three common sources for smear preparation?

1. Clinical Specimens, 2. Liquid/Broth Culture, 3. Solid Media Culture

How should a bacterial smear appear if properly prepared?

As a thin whitish layer or film where the print of a textbook (the newspaper test) is faintly readable through it.

What is the specific temperature and duration required for heat fixation on a slide warmer?

Warm at $65^{\circ}C$ for $10$ minutes.

What are the four primary purposes of fixation?

1. Preserves cell morphology, 2. Kills bacteria, 3. Attaches smear to slide, 4. Allows smear to take up staining material.

Which type of stain colors the background while leaving the cells unstained?

Negative Staining (Acidic Stain)

What is the cell arrangement where cocci are grouped in eights as two groups of four?

Sarcina

What is the difference in peptidoglycan thickness between Gram-positive and Gram-negative bacteria?

Gram-positive has thick peptidoglycan ($50\%$ - $90\%$ of cell wall), while Gram-negative has thin peptidoglycan ($10\%$ of cell wall).

List the four reagents used in the Gram stain procedure in order.

1. Primary Stain (Crystal Violet), 2. Mordant (Gram’s Iodine), 3. Decolorizer (Alcohol and/or Acetone), 4. Counterstain (Safranin)

Which genus of bacteria is known for having a high concentration of mycolic acid (long chain fatty acids) in its cell wall?

Mycobacterium

According to the Gram-staining general rules, which cocci are the exceptions (Gram-negative)?

Neisseria, Veilonelia, and Moraxella

What stain is used for Spirochetes since they are difficult to see with a standard Gram stain?

Silver stain

In the Gram Staining grading system, what does a grade of $4+$ indicate for bacteria?

$>25$ organisms per Oil Immersion Objective (OIO) field.

What is the difference between the Ziehl-Neelsen method and the Kinyoun method in acid-fast staining?

Ziehl-Neelsen uses heat/steam as a mordant, while Kinyoun uses a wetting agent called Tergitol.

In the Schaeffer-Fulton method (spore stain), what color do the spores and vegetative cells appear?

Spores appear green (Malachite Green), and vegetative cells appear red (Safranin).

Why should smears for capsule staining (Anthony Method) not be heat-fixed?

Heat causes cell shrinkage, which may create a false clear zone around the organism.

What reagent is used as both a decolorizer and a counterstain in the Anthony Method for capsule staining?

$20\%$ Copper Sulfate

Where should a slide be labeled and why?

The left end, because the right side is typically immersed in staining reagents during the process.