Microscopy Flashcards
What are the basic components of bright field microscopy?
- A light source to illuminate the specimen
- A condenser lens to optimise (center) illumination
- An objective lens for magnification
What is Kohler illumination?
It is a microscope technique that improves the imaging quality of microscopes by aligning and focusing the microscopes light source
how do you achieve Kohler illumination?
- Close the field diaphragm till you can see the sharp angles
- centre the small angled octagon
- re-open the field diaphragm
- take eyepieces off and adjust the light condenser until the barrel is completely filled with light
Magnification Vs Resolution
Resolution = Clarity or sharpness of an image
Magnification = The ability of a microscope to make an object appear smaller or larger than its actual size
What is optical resolution measured in?
Micrometres (µm)
How do you calculate the resolution of a lense?
R=0.61x λ/NA
Where:
R = Resolution
0.61 = A constant value
λ = Lambda (wavelength of light)
NA = Numerical aperture (NA = n x sin α)
How do you calculate Numerical Apeture
NA = n x sin α
Where:
NA = Numerical aperture
n = Refractive index
α = Half the angle of the lens aperture
How do you improve the resolution of a microscope?
- Decrease light wavelength
- Increase the Numerical Aperture
How do you alter the refractive index?
utilise specialty lenses designed to work in conjunction with water or oil
- Oil α = 1.5
- Water α = 1.3
How do you alter NA?
Decrease the focal length between the microscope and the tissue
What is an electron microscope
A microscope that utilises electron beams in place of light in tandem with electron lenses
What is a fluorescence microscope
Similar to a bright field microscope but uses a more intense light to excite fluorescent species in a sample
