BIOMG 4320 |Quiz 1 | Lectures 1-4| 2024
archaea *** Domain of unicellular prokaryotes that have cell walls that do not contain
peptidoglycan
bacteria *** single-celled organisms that lack a nucleus; prokaryotes
eukaryotes *** Cells that contain nuclei
main characteristics of eukaryotic cells that prokaryotic cells don't have *** - bound
organelles
- singular cellular membrane
- membrane bound nucleus -- DNA in nucleus
- complex membranous organelles (ER, Golgi, lysosomes)
- sexual reproduction requiring meiosis and fertilization
similarities between prokaryotic and eukaryotic cells *** - similar mechanisms for
transcription and translation
- genetic information encoded by DNA using same genetic code
- plasma membrane of similar composition and structure
resolution (D) *** the smallest distance between two points at whhich the points can be
recognized as two entities instead of one
formula for resolution *** D = 0.61 lamba/ N sin alpha
what is alpha in resolution equation *** the angular aperture, or half-angle, of the cone
of light entering the objective lens from the specimen
what is N in resolution equation *** the refractive index of the medium between the
specimen and objective lens
numerical aperture *** marked on objective lens -- defines range of how far system can
accept light
what is the limit of resolution of a light microscope *** - 200 nm
- depends on limitations in the values of alpha, lambda, and N
bright field image *** see how light goes through specimen
phase contrast image *** cells are surrounded by alternating dark and light bands, both
in-focus and out-of-focus details are simultaneously imaged in a phase contrast
microscope
, differential interference contrast (DIC) image *** -only narrow in-focus (optical slice)
region is imaged
- analogous to phase contrast microscopy with images looking 3D
what is fluorescence *** property of chemical where it absorbs light at one wavelength
(excitation) and emits light at a specific longer wavelength
--> in excitation and emission plots, emission plot is farther down right, showing that the
emitted photon has a longer wavelength
wide field fluorescence microscopy vs confocal fluorescence microscopy *** - confocal
utilizes pinhole to block out-of-focus light, leaving only the plane that is in focus to be
imaged.
- image serial optical sections, allowing for visualization of objects in the Z dimension as
well as X and Y
two types/light paths of confocal microscopy *** - point-scanning confocal microscope
--> pinhole excludes light from out-of-focus focal planes (takes focused images)
- spinning disk confocal microscope. --> good for recording dynamic events in live cells
deconvolution microscopy *** - uses calculated point-spread functions of out-of-focus
light to computationally remove fluorescence contributed by out-of-focus parts of the
sample
--> like confocal as it is a way to obtain sharp optical sections, but is largely
computational/intensive
total internal reflection fluorescence (TIRF) microscropy *** for imaging molecules
within a restricted focal plane near the coverslip
two-photon excitation microscopy *** for imaging deep tissue samples such as the
brain of a live mouse
light-sheet microscopy *** for rapidly imaging a large volume, for example, in
transparent living tissues
fluorescence recovery after photobleaching (FRAP) *** - reveals protein mobility
forster resonance energy transfer (FRET) *** - for studying protein-protein interactions
- uses two different fluorescent proteins so that when one is excited, energy will be
transferred to the second one by FRET, provided they are sufficiently close
super-resolution microscopy: localization microscopy *** - uses single-molecule
detection and localization
- image each fluorescent molecule individually and find the center of each spot, could
"beat" the resolution limit and separate spots that are too close for conventional
microscopy
archaea *** Domain of unicellular prokaryotes that have cell walls that do not contain
peptidoglycan
bacteria *** single-celled organisms that lack a nucleus; prokaryotes
eukaryotes *** Cells that contain nuclei
main characteristics of eukaryotic cells that prokaryotic cells don't have *** - bound
organelles
- singular cellular membrane
- membrane bound nucleus -- DNA in nucleus
- complex membranous organelles (ER, Golgi, lysosomes)
- sexual reproduction requiring meiosis and fertilization
similarities between prokaryotic and eukaryotic cells *** - similar mechanisms for
transcription and translation
- genetic information encoded by DNA using same genetic code
- plasma membrane of similar composition and structure
resolution (D) *** the smallest distance between two points at whhich the points can be
recognized as two entities instead of one
formula for resolution *** D = 0.61 lamba/ N sin alpha
what is alpha in resolution equation *** the angular aperture, or half-angle, of the cone
of light entering the objective lens from the specimen
what is N in resolution equation *** the refractive index of the medium between the
specimen and objective lens
numerical aperture *** marked on objective lens -- defines range of how far system can
accept light
what is the limit of resolution of a light microscope *** - 200 nm
- depends on limitations in the values of alpha, lambda, and N
bright field image *** see how light goes through specimen
phase contrast image *** cells are surrounded by alternating dark and light bands, both
in-focus and out-of-focus details are simultaneously imaged in a phase contrast
microscope
, differential interference contrast (DIC) image *** -only narrow in-focus (optical slice)
region is imaged
- analogous to phase contrast microscopy with images looking 3D
what is fluorescence *** property of chemical where it absorbs light at one wavelength
(excitation) and emits light at a specific longer wavelength
--> in excitation and emission plots, emission plot is farther down right, showing that the
emitted photon has a longer wavelength
wide field fluorescence microscopy vs confocal fluorescence microscopy *** - confocal
utilizes pinhole to block out-of-focus light, leaving only the plane that is in focus to be
imaged.
- image serial optical sections, allowing for visualization of objects in the Z dimension as
well as X and Y
two types/light paths of confocal microscopy *** - point-scanning confocal microscope
--> pinhole excludes light from out-of-focus focal planes (takes focused images)
- spinning disk confocal microscope. --> good for recording dynamic events in live cells
deconvolution microscopy *** - uses calculated point-spread functions of out-of-focus
light to computationally remove fluorescence contributed by out-of-focus parts of the
sample
--> like confocal as it is a way to obtain sharp optical sections, but is largely
computational/intensive
total internal reflection fluorescence (TIRF) microscropy *** for imaging molecules
within a restricted focal plane near the coverslip
two-photon excitation microscopy *** for imaging deep tissue samples such as the
brain of a live mouse
light-sheet microscopy *** for rapidly imaging a large volume, for example, in
transparent living tissues
fluorescence recovery after photobleaching (FRAP) *** - reveals protein mobility
forster resonance energy transfer (FRET) *** - for studying protein-protein interactions
- uses two different fluorescent proteins so that when one is excited, energy will be
transferred to the second one by FRET, provided they are sufficiently close
super-resolution microscopy: localization microscopy *** - uses single-molecule
detection and localization
- image each fluorescent molecule individually and find the center of each spot, could
"beat" the resolution limit and separate spots that are too close for conventional
microscopy
