Preparation of Specimens for Light Microscopy

Chapter 3

Why do cells and specimens need to be stained

Live unstained specimens have little contrast with the surrounding medium.

Live specimens are used to study what

Cell behavior

What does staining mean

Coloring microorganisms with dye that emphasizes certain structures

What is a smear

Thin film of a material containing microorganisms spread over a slide

When microorganisms are fixed (attached) to the slide what happens

Microorganism dies

Stains consist of positive and negative ions which are colored (chromophore) what are they called

Basic Dye and Acidic Dye

What is a basic dye

Chromophore is a cation (+)

What is a Acidic dye

Chromophore is an anion (-)

What is Negative Staining

Staining the background instead of the cell

What is a simple stain

Use of a single basic dye that highlights entire microorganism to visualize cell shapes and structures

What are examples of simple stains

Crystal Violet, Methylene Blue, and Safranin

What is used to hold the stain or coat the specimen to enlarge it

A mordant

What are differential stains used for

To distinguish between bacteria by using a gram stain or acid-fast stain

Name types of differential stains

Gram stain and Acid-Fast stain

What is a Gram Stain

Classifies bacteria into gram-positive or gram-negative

If bacteria is Gram Positive what does this mean

Has thick peptidoglycan cell walls

If bacteria is gram Negative what does this mean

Has thin peptidoglycan cell walls and layer of lipopolysaccharides

What is a Acid-Fast stain

Binds only to bacteria that has a waxy material in their cell walls, which is not decolorized by acid alcohol

What are Acid-Fast stains used for

Identification of Mycobacterium and Nocardia

What are special stains used for and give examples

Distinguished parts of microorganisms like

• Capsule staining

• Endospore staining

• Flagella staining

What are capsules

Gelatinous covering that doesn’t accept most dyes

How does negative staining for capsules work

Suspension of India ink or nigrosine contrasts the background with the capsule, which appears as a halo around the cell

What are Endospores

Resistant, dominant structures inside some cells that cannot be stained by ordinary methods,

When you stain an endospore, what color is the primary stain

Malachite green, usually with heat

When you stain an endospore, what color are the decolorized cells

water

When you stain an endospore, what color is the counterstain

Safranin

Endospores appear green within what type of cells

pink or red cells

What are flagellaS

Structures of locomotion

Flagella Staining uses what

A mordant and Carbolfuchsin