Lab quiz 1

Lab safety, Lab overview, Module 1, and Module 2 (except exercise 2-5)

What goes in the Biohazard bin?

1) live cultures on agar plates

2) Pipet tips

3) Serological pipettes

4) Gloves

5) Dropper (not the wrappers

6) Swabs (not the wrappers)

7) microcentrifuge tupes

What should be disposed in trash?

1) paper towels used for wiping benches

2) Gauze

3) wrappers from swabs, droppers, and serological pipettes

Compound brightfield microscope

Examines morphology (shape) of live microbial cells, or “fixed” specimen

Brightfield microscope

Specimen look dark against bright (white) background

• Since bacterial cells are transparent→ often have to stain cells before microscopy

Darkfield/ phase contrast microscope

• do not have to stain

• better contrast → species appear bright against dark background

• Examines cell morphology as well as motility in native state

Bacteria that is difficult to stain is examined using?

Darkfield or phase contrast microscope

Which microscope has the ability to provide most detailed image of species?

Phase contrast

• Can observe certain intracellular structures and inclusions

How does light reflect in brightfield vs darkfield?

Brightfield: Light passes through specimen, making cells appear darker

Darkfield: Special condenser ensures only light that is reflected off the specimen will enter objective

What special component does the phase contrast microscope have that brightfield and darkfield do not?

1) annular diaphragm- light transmitted is directed into cone shape

2) Phase-shifting element- shifts waves of refracted light

what does the compound microscope have as it’s component?

1) objective lens- first magnified by this lens (4x, 10x, 40x, 100x)

2) ocular lens- Further magnified 10x

Resolving power

Ability of the microscope to distinguish two objects (or points) positioned closely as two separate objects

• determined by wavelength of light used and the numerical aperture

Resolving power equation

wavelength of light used/ 2(numerical aperture of the objective)

Why must immersion oil be added?

Increase the amount of light going into the objective lens and obtain a clear image

• would be blurry otherwise

parfocality

Specimen remain in focus while switching from one objective to another

Brownian motion (not true motile)

When bacteria appear to just be vibrating in the same place instead of moving around

• result of random collisions of partle/cells

Current flow (not true motile)

Simultaneous movement, in one direction

• commonly observed when the slides are starting to dry out

Why do bacteria have a strong affinity for basic dyes?

Because their cell surfaces have an overall negative charge

Examples of basic dyes

1) Methylene blue

2) Crystal violet

3) Safranin

4) Malachite green

What are acidic dyes commonly used for

background staining

• stain everything but the cells

Simple stain

• One basic dye is applied to the specimen and all cells are colored in a nonspecific way

Differential stain

Successfully stain structural and cellular components of bacterial cells

• used to classify cell-wall composition, capsules, endospores, and flagella

Acid fast staining resisting gram-staining?

Due to the presence of a waxy layer on the surface of their cells

• are gram-positive

• Harsher staining method→ retains dye during the treatment

Capsules and slime layers

• Made up of polysaccharides

• Makes it easier to attach to surfaces and form biofilm

• Having capsules increase the pathogensis

Sporulation process

Results in the formation of specialized structures called endospores

• metabolically inactive and extremely resistant structures

A vegatative cell can produce _______

one endospore