Chapter 9 Microscopy

the ability to distinguish objects that are close together

Brightfield, Phase Contrast, Darkfield, DIC

view stained specimen with natural color

enhances contrast in unstained specimen, coverts invisible phase light to visible brightness

polarized light prisms to convert minute changes in specimen surface topography into intense 3D high contrast image

blocks direct light, light scattered at lower angles

produces base magnification, relays real image of object

collects light from light source, focuses light onto specimen

uses filter to visualize biomolecules, cells, tissues by making them glow

fluorescently tagging antibodies to visualize and locate specific proteins within cells + tissues

the primary antibody binds directly to the target antigen, while the secondary antibody recognizes and binds to the primary antibody. The secondary antibody is conjugated to a fluorophore, which generates a fluorescent signal that allows visualization of the target molecule

obtains high res and 3D images of fluorescently labeled specimen by eliminating out of focus light

proteins that emit light when exposed to UV/ blue light

illuminates and images fluorescent molecules within a thin layer near the cell’s surface (cell membrane)

technique used to monitor protein dynamics (movement), zaps proteins - flourescene goes away - if no recovery = no more of these proteins in this cell, recovery = cell trafficks new proteins in response - natural movement

detect if proteins in close proximity have a dynamic of interactions, 2 proteins use florophore (emission wavelength same) light on first proteins goes through and shines on second proteins which goes throug, if light comes out as expected there is indication that proteins interact

TEM & SEM

Takes sliced images of sample by shooting electrons through, uses slices to make a 3D model of sampleSca

provides deep surface view, high res, huge depth, actual surface view of sample