Chapter 2 Basic components of living systems Flashcards
Unit conversions
M –> MM –> um –> Nm. ×1000 for each forwards arrow
Magnification formula =
Image size/ actual size
Important points
- careful with units - need to be the same to cancel out
- try to avoid cm - stick to M, mm, um, nm
- choose appropriate decimal places/ significant figures
Scale bars
- length of scale bar measured = image size of scale bar
- Actual size = written
Magnification definition?
The number of times bigger an image is than the actual size of an object
Resolution definition
- the level of detail in an image
- the smallest distance 2 points can be apart, and still be resolved as separate points
Light microscope: max resolution & magnification
Max resolution: 200nm
Max Magnification: ×1500
Electron microscope: max mag & resol
Maz resolution is 0.5nm
Max mag is 500,000 - 3,000,000
What limits resolution?
Wavelength of energy used:
- Light energy: 400nm wavelength (average)
- Electron beam = 1nm
What is the defraction limit?
Resolution = 0.5 × wavelength
Uses of stains in microscopy?
- to increase contrast of an image
- making specific parts of the cell stand out
Fluorescent stains
- Commonly used with confocal microscopy, often combined with antibodies
- antibodies bind with specific molecules in the cell and stains emit light
Differential staining?
The use of 2 OR MORE STAINS AT ONCE
- Useful to identify cell types e.g Gram staining identifies 2 different types of bacteria - Gram positive + Gram negative
Prokaryotic
Pro - before
Prokaryotic - before nucleus
Eu-karyotic
True nucleus
Eukaryotic organisms are in these kingdoms:
Animalia
Plantae
Fungi
Protoctista
ALL in domain EUKARYA
Prokaryotic kingdom
Prokaryotae
2 domains:
- bacteria, archae
What is naked DNA
No histone
How can genetic material transfer across bacteria
The pilus (1 is pili)
What shape are bacterial chromosomes
Circular
Invagination definition
Infolding of membrane to increase SA
The human eye can distinguish objects about ??? apart
The human eye can distingush objects about 0.1mm apart - about the size of a human egg cell
Light microscopes how they work
- light is sent from a light source through a specimen, the image of which is magnified by glass lenses
Advantages of light microscopes
- cheap
- easy to use
- used to study living cells
Disadvantages of light microscopes
Light microscopes use visible light to create an image -> so resolution is limited to 200nm and magnification to ×2000
Laser scanning microscopes
- use a high power beam of light to create an image
- the laser passes over each single point ij the specimen and CREATES AN IMAGE over time
- 3D image
- more expensive than light microscopes have a high resolution
Electron microscopes
- Instead of beam of light like in traditional microscopes, electron microscopes use a beam of electrons to study specimens
Why are electron microscopes higher resolution?
Electrons have a higher wavelength and so can be used to create higher resolution images (micrographs)
What is a micrograph
A micrograph is a digital image taken through a microscope to show a magnified image of an object
2 types of em: SEM &TEM
- in both types a beam of electrons is focused by a magnet towards a specimen
- specimens must also be stained or coated with a metal and placed in a vaccum
- this preparation means electron microscopy can only be used to study dead specimens
SEM
- The electron beam hits the specimen and is scattered into a detector
- this means the microscope image can be 3D because the different orientations where electrons land are scattering in various directions
- 3d but low magnification and resolution
TEM
The electron beam travels through the specimen to a detector below
This allows for a higher resolution and magnification than SEM but a 2D image
SEM Vs TEM
Electron beam ✅
3D 〰️ 2D
MAX MAG - 200,000 〰️ 2,000,000
Max resolution- 20nm 〰️ 0.1nm
sample preparation - dry mount
solid specimens are viewed whole or cut into very thin pieces with a sharp blade - this is called sectioning
- the sample is then placed on the centre of the slide, and cover slip placed over
sample preparation -wet mount
specimens are suspended in a liquid such as water or an immersion oil. A cover slip is placed on from an angle
sample prep - squash slides
wet mount 1st prepared, then lense tissue is used to gently press down the cover slip. depending on the material, potential damage to a cover slip may be avoided by squashing the sample between 2 microscope slides
- good tech for soft samples
sample prep for smear slides
the edge of a slide is used to smear the sample, creating a thin even coating on another slide
-e.g used for blood sample
what is the cytosol?
aqueous interior of cells
differential staining can?
distinguish between 2 types of organisms that would otherwise be hard to identify
gram stain technique is used to?
seperate bacteria into 2 groups - Gram-positive and Gramnegative bacteria
acid fast technique is used for?
to differentiate species of Mycobacterium from other bacteria
4 stages of slide production?
- Fixing
- Sectioning
- Staining
- Mounting
what is fixing (stage 1)
- chemicals like formaldehyde are used to preserve specimens in as near natural state as poss
what is sectioning (stage 2)
- specimens are dehydrated with alcohols and then placed in a mould with wax or resin to form a hard block. This can then be sliced thinly with a knife called a MICROTOME
what is staining (stage 3)
- Specimens are often treated with multiple stains to show difv structures
what is mounting (stage 4)
- the specimens are then secured to a microscope slide and a cover slip placed on top
stains- risk management
many of the stains used in the preperation of slides are toxic or irritant. A risk assessment must be carried out before any practical work is started
magnification definition?
it’s how many times larger the image is than the actual size of the object being viewed