Cell theory & Microscopy Flashcards
Who first discovered cells?
Robert Koch
What is cell theory?
- All organisms composed of ONE OR MORE CELLS
- Cells are the SMALLEST UNITS of life - form basis of organisation (in all living things)
- Cells arise only by division of a PRE-EXISTING CELL
What can we do to understand a cells function?
Based on the COMPONENTS of the cell - VARIATION allows us to understand this
How many types of cells are in the human body?
At least 200 cell types
All have the same chromosomes
What is the Honey Fungus like?
Largest organism in the world (2,200 acres)
Probably <20 types of cells in this fungus
What is microscopy?
Produces enlarged image through magnification
Images can be sensor detected & computer manipulated/enhanced
What is resolution?
Ability to discriminate parts of an image
What are the two main types of microscopy used?
Light microscopy
Electron microscopy
What is light microscopy?
- Broad beam of illuminating light focussed on specimen by condenser lens
- Bright light passes thru img
Requires staining - often involves in cell death
What sort of section needs to be used for light microscopy?
Thin section
What is the limit of resolution and magnification for light microscopy?
Resolution = 0.2um
Which is 1000x
What are the 4 stages to prepare a section for light microscopy?
- Fixation
- Embed
- Section
- Staining
What is the purpose of fixation for light microscopy?
Stops the tissue falling apart, immobilises cells
(use fixative e.g. formaldehyde)
What is the purpose of embedding for light microscopy?
This provides mechanical support
What is the purpose of sectioning for light microscopy?
You need a thin section for light microscopy
Cut thin section using microtome (with metal, glass or diamond blade)
What is the purpose of staining for light microscopy?
Improves the visibility of the thin sections
What is the equation for magnification?
Total magnification = magnification of objective lens X magnification of eyepiece lens
What are the other types of light microscopy?
- Phase contrast
- DIC (Differential Interference Contrast)
- Fluorescence microscopy
- Laser scanning confocal light microscopy
What is phase contrast microscopy?
Increase ‘contrast’ of images using phase contrast microscopy, cellular components have different refractive indexes
(good for looking at components)
What is DIC microscopy?
Rates of change in refractive index = 3D shapes
(Differential Interference Contrast)
What is fluorescence microscopy?
Specimen stained w fluorescent dye or protein - UV light excites the specimens, makes high contract img
Contrasts to dark background
What protein is often used for fluorescence microscopy?
GFP from jellyfish (Green Fluorescent Protein)
What is laser scanning confocal light microscopy?
Imagining of a thin plane with a thicker specimen (non-sectioned)
Focussed laser beam excited fluorescent molecules in cells & tissues, scans at single points in planes (gives sharp images)
What is laser scanning confocal light microscopy useful for?
For living cells - can pool images to construct 3D model
What are the two types of electron microscopy?
- Transmission electron microscopy (TEM)
- Scanning electron microscopy (SEM)
How does electron microscopy work?
Uses a beam of electrons rather than light
Creates much more powerful images that light
How does electron microscopy work?
Uses a beam of electrons rather than light
Creates much more powerful images that light
How does TEM work?
Electrons pass through a very thin specimen - allows magnification of up to 1,000,000x
How must the specimen be prepared for TEM?
Fixed, dehydrated, sectioned & stained with electron dense metals (platinum or gold)
What happens to samples for TEM?
The samples could be rapidly frozen (cryofixatton)
Freeze-fracture reveals cell structure (knife used) cleaving cell then split surfaces using dense materials
How does SEM work?
Electron beam is scanned over the surface - electrons bounce back & strike a detector
What kind of image is produced by SEM?
Provides info on the surface details - appears 3D
Lower mag than TEM
What are the 3 main recent advances in microscopy?
- Cryo-electron microscopy
- Super resolution microscorpy
- Atomic force microscopy (AFM)
What is Cryo-electron microscopy?
Less harsh method, more intricate detail
Use deep frozen molecules in solution & gentler electron beams (these are collected on the other side of the cell)
Determines structure of biomolecules
What is super resolution microscopy?
Fluorescent based techniques
Overcomes traditional light resolution limit by various complex interference/tunneling methods
Allows resolution to ca. 100nm
What is Atomic Force Microscopy (AFM)?
Method to visualise surfaces at a molecular scale
uses fine pointed tip linked to cantilever arm, can move up & down as moves across surface
Detect fine movement by reflected laser beam
