SLIDE 1 Principles of Microscopy I: Point Spread Function and Resolution
Humberto Cabrera
Venezuelan Institute for Scientific Research International Centre for Theoretical Physics
Preparatory School to the Winter College on Optics: Advanced Optical Techniques for Bio-imaging
SLIDE 2
Wave optics view of the effect of the microscope lenses
SLIDE 3 Wave optics view of the effect of the microscope lenses
For a point object in a focal plane, what is the distribution of light at and near image plane (PSF)?
SLIDE 4 Huygens wavelet approach
- Each of the infinite points in the
emerging wavefront acts like a point source
- Each point emits a wavelet
- All wavelets from the same wave front
are mutually coherents
- That is, they oscillate synchrony
- Therefore, they interfere wich each
- ther in a predictable way
SLIDE 5
Interference of wavelets
SLIDE 6
Effect of two wavelets
SLIDE 7
Interference from two wavelets
SLIDE 8
Numerical aperture and resolution
SLIDE 9
Numerical aperture and Resolution
SLIDE 10
Numerical aperture and Resolution
SLIDE 11
Wavelength and Resolution
SLIDE 12
Effect of multiple wavelets
SLIDE 13
Effect of three wavelets
SLIDE 14
Effect of five wavelets
SLIDE 15
Effect of nine wavelets
SLIDE 16
Effect of all the wavelets: the PSF
SLIDE 17
Effect of the numerical aperture on fringes
SLIDE 18
PSF is smaller for shorter wavelength light
SLIDE 19
Effect of multiple wavelets: Airy discs
SLIDE 20
Airy discs parameters
SLIDE 21
PSF light distribution near the image plane (xy and xz)
SLIDE 22
Numerical aperture´s influence on PSF: bigger effect on axial than lateral spread
SLIDE 23
Resolution in the light microscope
SLIDE 24
Rayleigh criterion (lateral and depht)
SLIDE 25 Low NA Medium NA High NA
The higher the NA the smaller the Rayleigh criterion and the better the resolution
SLIDE 26
Resolution limit
SLIDE 27
Resolution: the sampling theorem
SLIDE 28
Resolution: the sampling theorem
SLIDE 29
Resolution limit: the sampling theorem
SLIDE 30
SLIDE 31
SLIDE 32
Thanks
SLIDE 33
I would like to thank Professor Jeff Lichtman (Harvard University) (iBiology.org microscopy course and educator resources) for his useful figures, lectures and remarks which we used for the preparation of the lecture
