2.1.1 MICROSCOPY Flashcards

1
Q

magnification

A

the amount of times larger the image of an object is compared to the objects actual size

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2
Q

resolution

A

how well the microscope can distinguish between two separate points on an image as two separate objects

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3
Q

how does an optical (light) microscope work

A
  • light passes through thin sample layer that is mounted on a glass slide
  • light is focused through lenses forming an image for viewing on eyepiece
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4
Q

pros of optical (light) microscope

A
  • cheap
  • easy to use
  • portable
  • able to study living specimen/ tissue
  • can produce colored images
  • can view basic structures (e.g: nucleus)
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5
Q

cons of optical (light) microscope

A
  • low resolution
  • low magnification
  • specimen may require staining before viewing
  • cannot be used in places with bright light
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6
Q

why does optical (light) microscopes have low resolution

A
  • the wavelength of light is too long to pass through two separate structures
  • so it cant distinguish between two close-together but separate objects in the image
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7
Q

magnification (light microscope) equation

A

objective lens x magnifying lens

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8
Q

photomicrograph

A

image seen viewing specimen on a light microsocope

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9
Q

Laser scanning confocal microscope LSCM

A

uses laser light to scan object point by point and assemble a pixel image on a computer screen

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10
Q

pros of LCSM microscope

A
  • can view image in different dimensional images
  • high resolution
  • viewing of living things
  • has depth selectivity to focus on organelles at different depths
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11
Q

how do a transmission electron microscope work

A
  • electron gun fires electron beam down
  • condenser lens focuses electrons in one beam
  • beam passes through chemically-fixed, dehydrated, and stained specimen
  • objective lens focuses beam into image
    image enlarges and viewed on phosphor screen
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12
Q

pros of TEM

A
  • very high magnification allowing viewing of cells in great depth
  • high resolution of a samples inner structure
  • 2d image of cells ultrastructure
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13
Q

cons of TEM

A
  • sample needs to be thinned and chemically fixed, and dehydrated
  • shows 3d structures in 2d which can be difficult to interpret
  • difficult to use
  • cant view live specimens
  • requires training as difficult to use
  • large and expensive
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14
Q

how is specimen prepared for TEM

A
  • specimen is fixed in formaldehyde to preserve cells structure
  • dehydrated as water from cell is replaced with organic solvent (ethanol)
  • replace solvent w resin
  • thin down specimen
  • stain using metal salts to increase contrast
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15
Q

how does a scanning electron microscope work SEM

A
  • electron gun shoots electron beam
  • electrons reflect off sample surface
  • the reflected electrons are focused on screen
  • producing a 3d image of cells
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16
Q

pros of scanning electron

A
  • produces 3d visual of cells topography
  • fast (produces visual in under 5 mins)
  • high magnification
  • high resolution
17
Q

cons of SEM

A
  • expensive
  • require training to operate
  • risk of radiation exposure
  • cannot view live cells
  • uses harmful metallic salt stains
18
Q

preparing specimen for SEM

A
  • fix sample using formaldehyde
  • dehydrate sample using solvent (ethanol) to replace water
  • mount sample on Aluminium stub
  • coat sample in conductive metal (gold)
19
Q

SEM vs TEM

A

SEM = 3d vs TEM = 2d

SEM = cell topography vs TEM = ultrastructure inside cell

SEM = thick sample vs TEM = thin sample

SEM = lower resolution vs TEM = higher resolution

SEM = lower magnification vs TEM = Higher magnification

20
Q

methylene blue

A

all purpose stain

21
Q

acetic orcein

A

binds to DNA and stains chromosomes dark red

22
Q

eosin

A

stains cytoplasm

23
Q

sudan red

A

stains lipids

24
Q

iodine in potassium iodide

A

stains cellulose in plant cell walls yellow
and starch granules blue/black

25
equation for magnification (mia)
magnification = image / actual
26
how to draw cell
draw cell in pencil label structures give magnification scale
27
eyepiece graticules
measuring device in eyepiece of microscope
28
stage graticule
scale placed on microscope stage to calibrate eyepiece graticules
29
differential staining
Specimens stained with multiple dyes so different tissues in specimen show
30
onion cell microscope experiment
use scalpel to peel off thin onion epidermis (specimen) lay specimen flat on glass slide using tweezers add two drops of iodine in potassium iodide to stain (eg: cell wall is now dyed yellow) apply cover slip on top at 45 degree angle clip slide on to microscope stage for viewing
31
cheek cell microscope experiment
collect epithelial cells by rubbing sterile cotton swab on inner cheek smear cells onto clean microscope slide add two drops of methylene blue dye and let stain for 30 seconds place cover slip on top at 45 degree angle clip slide onto microscope stage for veiwing
32
pond water microscope experiment
using pipette collect a few drops of pondwater place one drop in middle of clean microscope slide add methyl cellulose: slow organisms from moving add stain (methylene blue or I in KI) place cover slip on top at 45 degree angle clip slide onto microscope stage
33
blood microscope experiment
prick finger with needle dropping one drop of blood onto edge of clean microscope slide at 45 degree angle hold slide and spread drop across slide let blood air dry and add giemsa stain wait 2 minutes add drops of distilled water and let dry place coverslip on and clip slide onto stage
34
artefacts (microscopy)
structures that appear in an image but are not part of the specimen