Chapter 2- Basics of Living Systems Flashcards
How do you prepare a dry mount in sample prep?
- Solid specimens are either viewed whole or cut into thin slices (sectioning)
- Specimen is placed on the centre of slide and a coverslip is placed over a sample.
How do you prepare wet mount in sample prep?
- specimens are suspended in liquid e.g. water/immersion oil
- cover slip is placed on from an angle (45º)
- e.g. aquatic sample + living organisms
How do you prepare squash slides in sample prep?
- wet mount is first prepared
- lens tissue is used to gently press down cover slip
- use 2 microscope slides to avoid damage to cover slip
- good for soft sample
- e.g. root tip for cell division
How do you prepare a smear slides in sample prep?
- edge of slide is used to smear sample
- creates thin, even coating
- cover slip is then placed over sample
- e.g. blood smear slide.
Why do we use staining?
- The cytosol (aqueous interior) of cells and other cell structures are often transparent
- Stains increase contrast (different components in cell take stains to different degrees.)
- The increase in contrast allows components to become visible so they can be identified.
How do positively charged dyes like “Crystal Violet” and “Methylene Blue” work?
-The positively charged dye is attracted to negatively charged materials in cytoplasm, leading to staining of cell components.
How do negatively charged dyes like “Nigrosin” or “Congo Red” work?
The negatively charged dye is repelled by the negatively charged cytosol.
These dyes stay outside cells, leaving the cells unstained, which then stand out against the stained background.
=Negative Stain Technique.
What is Fixing, Sectioning, Staining and Mounting in sample prep?
Fixing- chemicals like formaldehyde are used to preserve specimens in as near-natural a state as possible
Sectioning- specimens are dehydrated with alcohols and then placed in a mould with wax or resin to form a hard block. This can be sliced thinly with a microtome.
Staining- specimens are often treated with multiple stains to show different structures.
Mounting- the specimens are then secured to a microscope slide and a cover slip placed on top.
Describe what magnification is briefly:
Magnification is how many times larger the image is than the actual size of the object being viewed.
What is resolution in microscopy?
Resolution is the ability to see individual objects as separate entities.
It is limited by the diffraction of light, as it passes through samples.
What is the calculation for magnification?
Magnification = image size / actual size
What is an eyepiece graticule?
- An eyepiece graticule is a glass disc marked with a fine scale of 1 to 100.
- The scale has no units and remains unchanged whichever objective lens is in place.
- The relative size of the divisions increases with each increase in magnification.
- The scale on the graticule at each magnification is calibrated using a stage micrometer.
What is a stage micrometer?
- A stage micrometer is a microscope slide with a very accurate scale in μm.
- the scale marked in the micrometer slide is usually 100 divisions = 1nm, so 1 division = 10 μm.
How does a Transmission Electron Microscope (TEM) work?
In a transmission electron microscope, a beam of electrons is transmitted through a specimen and focused to produce an image.
Resolution of 0.5 nm
How does a Scanning Electron Microscope work?
In a scanning electron microscope, a beam of electrons is sent across the surface of a specimen and the reflected electrons are collected.
Resolution = 3-10 nm
