Module 2 Flashcards
(167 cards)
light microscope Advantages
• Wide range of specimens can be observed • Specimens can be alive • Specimens can be whole, or embedded in wax ten sectioned
light microscope Disadvantages
• Non-coloured specimens must be stained for specific organelles or molecules • Relatively low resolution does not give detailed information
light microscope Magnification
• Up to 1500x in total
Resolution
• 200nm • Limited by the wavelength of visible light
light microscope Specimen Preparation • into a thin slice
Staining ○ Applying a coloured stain to the sample which binds to certain chemicals/structures, improving their visibility ○ Acetic orcein stains DNA dark red ○ Gentian violet stains bacterial cell walls
Sectioning ○ Specimen is embedded in wax to preserve structure of sample cell walls while cutting them
To correctly use a light microscope
you also need to calibrate the eye piece graticule using the stage micrometer
— this step allows you to measure the size of cells and structures that you are observing.
how to calibrate the eyepiece graticules
To do this, align the stage micrometer (a microscope slide with a scale on it) with the eye piece graticule, then use the reading from the scales to calculate the calibration factor from the objective lens.
Using A Transmission Electron Microscope (TEM)
emits an electron beam through a very thin prepared sample. Electrons penetrate the denser parts of the sample with greater difficulty and this gives the contrast in the 2D image produced.
Using A A Scanning Electron Microscope (SEM)
emits an electron beam directly onto a sample such that none of the electrons penetrate it. Instead, they ‘bounce off’ the sample and are received on asensor, producing a 3D image.
A Scanning Electron Microscope (SEM) Magnification
• SEM: Up to x100, 000
Using A Transmission Electron Microscope (TEM) Magnification
TEM: Up to x500, 000
Resolution (TEM)
• 0.5nm 2000x times more than light microscope • Produces detailed images
resolution SEM)
3-10nm
Magnification
• The degree to which the size of an image is larger than the object itself
Resolution
• The degree to which it is possible to distinguish between two points on an object that are very close together • The higher the resolution, the greater the detail that you can see
Specimen Preparation em
• Specimen needs to be prepared correctly
• Fixed to make it firm
• Dehydrated and embedded in resin
• Stained using metal salts or metal particles
• Mounted on a copper grid
• Placed in a vacuum
• Staining
○ Specimens are stained with metal salts or particles ○ This causes electrons to scatter differently, giving contrast
Advantages
- Produces detailed images of the structures inside cells
* SEM produces detailed 3D images showing contour of cells
Disadvantage
• Electron beams deflected in air, so sample must be in a vacuum • Samples must be dead • Extremely expensive • Large piece of equipment • Use requires a high degree of skill and training
Structure Nucleus
• Largest organelle • Surrounded by nuclear envelope • Contain chromatin • Nucleolus at the centre
Structure Nucleolus •
• Dense spherical structure inside nucleus
Structure Nuclear Envelope
• Surrounds nucleus • 2 membranes with fluid between them • Nuclear pores go through envelope
Structure Rough Endoplasmic Reticulum
• Flattened membrane sacs called cisternae • Continuous with outer nuclear membrane • Studded with ribosomes
Structure Smooth Endoplasmic Reticulum
• Flattened membrane sacs called cisternae • Continuous with outer nuclear membrane • (No ribosomes)
Structure Golgi Apparatus
• Stack of membrane bound, flattened sacs