Diagnostic Techniques and Applications Flashcards
What is the Rayleigh Criterion?
Two points on an object are distinguishable if the central maximum from one is beyond the first minimum of the second.
Principle of conventional microscopy?
The sample is uniformly illuminated. Dyes or stains are then applied to add contrast to specific areas of interest.
The final image is virtual and magnified.
Minimum separation distance for conventional microscopy?
dmin = λ/NA
(2NA) for confocal microscopy
Fluorescence microscopy?
Uses UV rather than visible light. Gives info on different types of material with fluorescence.
What is the Stokes’ shift, visible in fluorescence microscopy?
A decrease in photon energy from to absorption/reemission at a lower energy. Caused by the electron de-exciting to a higher energy level than before.
Benefits of using FRET imaging?
It has a resolution ~10x lower than typical diffraction limited microscopes so can technically break the diffraction limit. It is used as a ‘microscopic ruler’ due to the ‘Förster distance’ being known at 10nm.
Principle of FLIM imaging?
Laser scanned across the sample, exciting molecules and causing fluorescence. Fluorescence light is recorded with a photomultiplier tube and constructs an image of the sample.
Multi-photon imaging?
Pulses of low energy photons cause a buildup of energy that eventually leads to fluorescence. This creates a focused image of a higher resolution than FLIM.
Principle of pulse oximetry?
- A red and an infrared light source are shone through a finger.
- The amount they are attenuated by the blood is measured on the other side.
- Oxygenated and deoxygenated haemoglobin have very different absorption coefficients.
- Higher absorption also indicates pulse due to increased blood volume.
Equation for blood oxygenation?
SO2 = HbO2/(HbO2 + Hb)
Isobetic point in spectroscopy?
A point where absorbance is the same for both oxygenated and deoxygenated haemoglobin, so absorption curves cross.
What is spectroscopy?
Using a white light source to obtain an absorption spectrum across a set of wavelengths.
Multispectral imaging?
Illuminate a large area, then image through several different filters to build a composite image.
What is the photoacoustic technique?
- Tissue is illuminated with a ns pulsed laser.
- Preferential absorption by a λ dependent chromophore eg Hb.
- Excitation and thermal expansion of the tissue emits ultrasonic sound waves.
- These can be detected by a transducer.
Conditions for the input light in photoacoustic imaging?
The laser pulse length must be less than the thermal and acoustic relaxation times (τr & τrs)
Eg ~ns pulses
NIR wavelength (~800nm)
Low energy (~30mJ)
