Luminex 100 User Manual Version 1.7 - Manual (Page 13)

x MAP Technology Introduction
PN 89-00002-00-063 Rev. A 1 - 3
Excitation
The excitation system in the Luminex 100 analyzer uses two solid-
state lasers. A reporter laser excites fluorescent molecules bound to
biological reactants at the microsphere surface, and a classification
laser excites fluorochromes embedded in the microsphere. The lasers
illuminate the xMAP microspheres as they flow single-file through
the cuvette. These fluorescent signals are discriminated with
selective emission filters and are converted into intensity units by a
digital signal processor.
xMAP Microspheres
The xMAP microspheres are highly uniform, polystyrene particles
that have been crosslinked during polymerization for great physical
and thermal stability. Varying amounts of fluorochromes embedded
within each microsphere give each microsphere set an unique
fluorescent signal. To ensure the stability of this address, it is
essential to protect the xMAP microspheres from light. Do not
subject xMAP microspheres to prolonged high temperatures, and
protect them from freeze/thaw manipulations.
Reporter Fluorochromes
Each xMAP microsphere is dyed to emit light in a certain
classification channel. All xMAP microspheres of a given emission
represent a distinct assay within a multiplex of assays. A reporter
channel is used to detect fluorescence bound to the surface of each
microsphere, and each reporter emission quantitates each of the
distinct assays. Only one reporter emission is used for multi-analyte
assays in the Luminex 100 analyzer. The Luminex 100 analyzer uses
an excitation wavelength of 532 nm, and has an emission wavelength
of 575 nm
± 12 nm.
Fluorochromes R-Phycoerythrin, and Alexa
®
532 work well with the
Luminex 100 analyzer, but you are not limited to these two. You may
have success using other fluorochromes such as Cyanine 3.
R-
Phycoerythrin
Alexa
532
Formula weight (Daltons) 240,000 470
Absorbance max (nm) 480, 546, 565 531
Extinction max (M
-1
cm
-1
)
1,960,000 83,800
Emission max (nm) 578 554
Quantum yield 0.82 0.8