1 Cells and microscopy Flashcards
how is magnification calculated?
image size / actual size
how is total magnification calculated?
eye piece magnification x objective magnification
what are the differences between light and electron microscopes?
light v electrons
light uses living; electron has vacuum ∴ dead samples
light can distinguish colour; electron can’t
light has low mag. and res.; electron has high mag. and res.
what does an electron microscope use to focus?
an electromagnet
what is the maximum resolution for a
- light microscope
- electron microscope?
- 200nm
- 0.1nm
what is the maximum magnification for a
- light microscope
- scanning electron microscope (SEM)
- transmission electron microscope (TEM)?
- 1500x
- 100,000x
- 500,000x
what is the definition of magnification?
the degree of enlargement of an image to reveal further detail
what is the definition of resolution?
the ability to distinguish between two different points and to see detail
what is the use of a microtome?
to cut extremely thin non-living specimens
how does a TEM produce an image?
e- passed through specimen and scattered
magnetic lenses focus image on fluorescent screen/photographic plate
how does an SEM produce an image?
e- knock other e- from surface of specimen
what is added to TEM specimens?
heavy metal impregnation
what is added to SEM specimens?
coated with carbon/gold
outline the process of temporary staining
fixation
staining
mounting
outline the process of permanent staining
fixation (immersion in gluteraldehyde)
dehydration (incr. alcohol content)
clearing (xylol removes alcohol)
embedding (in epoxy/resin)
sectioning
staining
mounting
what is cryosectioning?
tissue embedded in gel medium is rapidly frozen to -20/-30.C
cut with cryostat into 5-10 um sections, then stained
what is the purpose of differential staining?
to make certain structures appear darker or different in colour from other structures
how is Leishman’s stain applied?
blood smear prepared
fixed with methanol for 2 mins
distilled with water for 6 mins
slide washed until pink to naked eye
how is Wright’s stain applied?
blood smear prepared
dipped in stain for 15-30s
dipped in water for 15-45s
dipped in fresh water for 25s
dried vertically
oil applied
what is the purpose of a blood smear?
to observe the appearance of the blood:
- presence/absence of cells
- cell morphology
- cell health
how is a blood smear achieved?
place a small drop of blood at the end of a clean, sterile, dry slide
another slide used to spread blood at 30. angle
immediately labelled
slide left to dry
fixative used to preserve cells
what are the functions of the blood?
delivery of O(2) and nutrients to tissue
removal of waste products from tissues
immunological protection
clotting
transport of cell-signalling molecules
buffer to regulate body pH and temperature
what is the function of erythrocytes?
{red blood cells, RBCs}
to deliver oxygen to tissues as oxyhaemoglobin
how are erythrocytes produced?
produced from erythropoietic stem cells in bone marrow following stimulation from erythropoietin
how is the structure of an erythrocyte related to its function?
no nucleus; few organelles (incl. mitochondria)
biconcave (incr. SA)
production of haemoglobin (for oxygen transportation)
how is the structure of a neutrophil related to its function?
lobed nucleus
granular cytoplasm
many lysosomes containing hydrolytic enzymes (phagocytosis)
what are the requirements of a haemocytometery sample?
mixed (representative)
known dilution (accurate counting)
how is a cell count performed using a haemocytometer?
count number of cells in 3-line square using NW rule = ___
find volume (0.2 x 0.2 x 0.1 = 0.004 mm3)
in 1 mm3 = 1/0.004 x ___ = ~~~ cells
account for dilution = ~~~ x dilution factor = …….. cells