Flow cytometry
Principle
Basic
instrumentation
Advantages
Disadvantages
Measurement of multiple
physical characteristics of
a single cell
fluidic system, laser(s),
optics, and
electronics/external
computer system
-High speed analyses
-Measures single cells and
a large number of cells
- Simultaneous analysis
multiple parameters
- Quantification of
flurescence intensities
-Sorting of predefined
cells populations
-Portable equipments
-Very expensive and
sophisticated
instruments;
-Requiries management
by a highly trained
specialist and on-going
maintenance by service
engineers.
-Complex instruments are
prone to problems with
the microfluidics system
(blockages) and also
require warm-up, laser
calibration and cleaning
for each use.
-Needs single cell
particule;
-Tissue structure is lost;
-Litle information on
intra-celular distributions
Ratio-referencing
fluorometry
Measurement of emitted
fluorescent light
power supply, xenon
source, excitation
monochromator, cell,
emission monochromator,
detectors, excitation
signal amplifier, emission
signal amplifier
-Fluorescence intensity
measurements are
more sensitive than
absorbance
- Rapid scanning to get
exitation & emission
spectrum
-“Inner filter effect”
-Concentration quenching
-Light scattering
-Effects exerted by cuvet
material and solvent
-Background fluorescence
from sample matrix
-Effects of temperature
fluctuations
-Photodecomposition
Nephelometry
Measurement of the light
scattered by a particulate
solution
Light source, collimating
optics, sample cell,
collection optics,
comprising light ,
scattering optics, detector
filter, and detector
-Able to measure the
concentration of a
solution that
contains particles too
large for absorption
spectroscopy
- Very rapid procedure
-Simplicity in
measurement
-Accurate
-antigen excess
-Influenced by qualities of
matrix
-High cost
-Easily damaged
-Require high power
supply