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Luminescent Processes
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Chemiluminescence is the emission of light from a chemical reaction that
occurs at or near ambient temperatures. A vast number of reactions give
rise to the emission of light in solution, but only a few have
sufficiently high efficiencies of chemiluminescence to have been used
for analytical purposes. Bioluminescence is a luminescent process
mediated by an enzyme or other biological system. For further
information on the vast array of chemical reactions that produce light
please refer to excellent reviews by McCapra and Beheshti in "Bioluminescence
and Chemiluminescence: Instruments and Applications", 1985, Ed. K. Van
Dyke, CRC Press, Boca Raton, FL. pgs. 9-42; and by Schuster and Smith,
"Adv. Phys. Org. Chem.", 1982, 18, 187.
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| Chemi- and bioluminescent systems can either be of the "glow" type,
where the emission of light builds slowly and reaches a maximum after a
substantial incubation time, typically minutes or hours, or of the
"flash" type where the addition of reagent(s) causes the immediate
emission of light, typically over milliseconds or seconds. An example of
glow reactions are those generated by enzyme systems such as alkaline
phosphatase using dioxetane phosphates as substrates. A typical flash
reaction is generated with an Acridinium ester.
Flash type systems such as the Acridinium esters have high or
moderate chemiluminescence efficiencies. The chemistry of this light
production is well understood. The simple triggering conditions
contribute little to the background signal and are an added benefit. At
the end of any immunological or nucleic acid binding reaction, signal
generation is immediate and unaffected by temperature variations. These
advantages have lead to the use of flash reactions in rapid quantitative
detection applications. The small molecular weight luminescent molecules
are easily attached via NHS or isothiocyanate chemistry and have minimal
effects on immunological or nucleic acid binding properties.
Glow type systems are excellent for qualitative systems such as
identification of proteins on gels, or for quantitative systems such as
immunoassays if sufficient temperature control can be maintained. For
the detection of proteins and nucleic acids on gels, the gel needs only
to be developed with enzyme labeled probe, bathed in an appropriate
substrate, and placed against a photographic film. After a brief
exposure the film is developed and labeled material can be visualized.
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There has been extensive use of chemi- and bioluminescent systems for in
vitro diagnostics over the past decade. The primary use of luminescence
in research has been molecular biology applications for the detection of
proteins and nucleic acids on gels, and in the visualization of
expressed proteins in cells. We would suggest you contact companies that
sell high quality products for applications such as Western blotting
visualization. With the exception of our Correlate-CLIA immunoassays,
there are no companies selling small molecule luminescent immunoassays,
such as PGE2, PGF2alpha, etc. We also sell kits
that utilize chemiluminescence for the detection of other biological
molecules at very low concentrations.
Because of the diversity of chemi- and bioluminescent molecules and
systems, it is suggested that the information about the individual
compounds or kits be reviewed for complete details. Please see the
complete line of our chemiluminescent molecules for further applications.
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| One important consideration with luminescent systems is the choice
of suitable detection devices and equipment. With gel based luminescent
detection systems, the only necessary component is a piece of suitable
photographic film. With quantitative detection systems, the choice of an
instruments with the required dynamic range, sensitivity,
reproducibility of injection, temperature control, on-board software for
data analysis, and other factors must be considered. For flash type
systems we suggest a photon counting luminometer that uses a syringe or
bellows pump type reagent injector. These types of injectors have the
best reproducibility for injection function. |
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