Microbiology Chapter 3: Microscopy and Staining

Vocabulary terms covering unit conversions, types of microscopy (Light, TEM, SEM), slide preparation, and staining techniques from Chapter 3 of the lecture notes.

Measurement conversion: Small to Large

To convert from a smaller unit to a larger unit, divide by $1000$.

Measurement conversion: Large to Small

To convert from a larger unit to a smaller unit, multiply by $1000$.

Total magnification

Calculated by multiplying the magnification of the objective lens by the magnification of the Ocular lens.

Immersion oil

A substance used with the oil immersion lens to reduce light loss between the slide and the lens.

Fluorescence microscopy

A technique where specimens are stained with fluorochromes and viewed through an ultraviolet light source, making microorganisms appear as bright objects against a dark background.

Immunoassays

A process where an Antibody is designed to bind to a certain target; if the target is present, the antibody attaches to it and produces a measurable signal indicating presence and quantity.

TEM (Transmission Electron Microscope)

Produces electron micrographs of thin sections of organisms with a magnification of $10,000 \times$ to $10,000,000 \times$ and a resolving power of $10\,pm$.

SEM (Scanning Electron Microscope)

Provides three dimensional views of the surfaces of whole microorganisms with a magnification of $1000 \times$ to $500,000 \times$ and a resolution of $0.Sum$.

Smear preparation steps

1. Place small sample on slide; 2. Spread into a thin film; 3. Allow to air-dry completely; 4. Fix the smear by passing the slide through a Bunsen Burner Flame several times.

Fixing

A process that kills microorganisms, attaches them to the slide, and preserves their natural state with minimal distortion.

Basic dye

A positively charged (cationic) dye that binds to negatively charged components of bacterial cells (e.g., crystal violet, methylene blue, Safranin).

Acidic dyes

A class of stains that carry a net negative charge and are used primarily for negative staining (e.g., eosin, acid fuchsin, nigrosine).

Simple stains

An aqueous or alcohol solution of a single basic dye used to highlight the entire organism.

Mordant

A substance used to improve the bonding between the stain and the specimen.

Differential stains

Staining methods such as the gram stain or acid-fast stain used to distinguish between different types of bacteria.

Gram-positive color

Microorganisms that appear purple after the staining process.

Gram-negative color sequence

Bacteria appear purple until the decolorizing agent is applied, becoming colorless, and then turn pink/Red after the counterstain.

Acid fast stains

Stains that bind only to bacteria with a waxy material in their cell walls that is not decolorized by acid-alcohol; used to identify Mycobacterium.