Microscopy

Magnification

Increase in size

Resolution

The ability to see two different points as separate

Contrast

Different in light intensity or color between a specimen and its background

Refraction

When light changes direction upon entering a medium

Refractive index

Degree to which a material slows transmission speed

Concave lens

Used to redirect light path in microscopes

Convex lens

Used for focusing light

Compound light microscope features

Uses visible light to illuminate cells using two sets of lenses (ocular and objective)

How do you calculate total magnification?

Ocular lens x objective lens

What is another name for compound light microscopes?

Brightfield

Limitations of brightfield

Needs contrast to visualize cells and need to use oil immersion

What does oil immersion do?

Prevents refraction of light away from the lens to improve resolution at high magnification

Phase contrast microscopy

Produces dark cells on light background, best for visualizing live cells

Darkfield microscopy

Cells appear light on a dark background, best for high contrast, high resolution

Fluorescence microscopy

Visualize cells that emit light (fluoresce), cells glow on dark background, best for clinical diagnostics

Autofluoresce

Emit light on their own

DAPI

Fluorescent dye

Immunofluorescence

Uses antibodies to visualize specific proteins

Direct immunofluroescence

Fluorochrome attaches to antibody which attaches to antigen (target protein)

Indirect immunofluorescence

Secondary antibody with fluorochrome attaches to primary antibody which attaches to antigen (target protein)

Differential interference contrast (DIC) microscopy

Best for visualizing cell structures, like the nucleus, in 3D

Confocal scanning laser microscopy (CSLM)

Scans layers of sample to make 3D image, good for generating images of thick samples like biofilms

Electron microscopy

Uses electrons instead of visible light

Transmission electron microscopy (TEM)

Best for visualizing molecular structures or viruses (ex: proteins). Sample must be thin and stained to visualize

Scanning electron microscopy (SEM)

Uses heavy metal staining to produce an image. Best for visualizing the surface only