Luminex xMap® Technology
The Luminex System, which is based on Luminex's xMAP® technology, combines flow cytometry, fluorescent microspheres, and traditional biochemistry into a flexible, multi-analyte profiling technology. It delivers highly sensitive, accurate and cost-effective results for the diagnostic clinical laboratory. Over the last 10 years, Luminex's xMAP technology has found widespread usage in both protein and nucleic acid-based detection assays for a broad range of clinical testing areas (e.g. as allergic disease, biomarker identification, pharmacogenomics, infectious disease, cystic fibrosis (CF) testing, genetic testing, and immunological monitoring). The core xMAP technologies that make up the Luminex System are the xMAP microspheres and the Luminex analyzer. The xMAP microspheres are a family of 100 fluorescently dyed 5.6 micron polystyrene microspheres, which serves both as a spectral identifier and a solid surface to build analyte-specific assays (Figure 1).
The xMAP detection system can identify to which set the microsphere belongs based on the unique spectral signature afforded by internally filling each of the 100 bead sets with a known ratio of red and infrared fluorophores. These color-coded microspheres are then conjugated to reagents, (e.g., antibodies, receptors, peptides, oligonucleotides, or streptavidin) specific to a particular bioassay, allowing the capture and detection of up to 100 specific analytes from a heterogeneous sample.
The Luminex analyzer is a flow cytometry-based instrument that combines fluidics, 2 lasers, 4 detectors, and real-time digital signal processing to distinguish the 100 different sets of color-coded microspheres, each bearing an analyte-specific assay. The fluidics system of the Luminex analyzer aligns the microspheres into single file as they enter a stream of sheath fluid and then enter a flow cell (Figure 2).
Once the beads are in single file within the flow cell, each bead is individually interrogated by the red 635 nm laser which is used to excite the dyes inside the beads to determine their "color" or "region" and also to discriminate doublets by light scatter. The green 532 nm assay laser is used to excite phycoerythrin (PE) which is the reporter dye captured during the assay (Figure 3).
The fluorescent signal generated from the lasers exciting both the dyes within the microspheres and the PE reporter dye molecules are distinguished by 4 detectors within the Luminex analyzer (Figure 4). One detector measures side scatter to size the microspheres and remove signals related to other materials passing through (typically air bubbles or microsphere aggregates). A second measures the internal red dye concentration, while the third measures the internal inferred dye concentration. The integration of these data defines and identifies the bead. A photomultiplier tube (PMT) associated with the green laser then measures the reporter signal for the event based on size and dye content. Depending on the specific protein or nucleic acid-based bioassay, the reporter signal intensity can be used to either qualitatively identify a biomarker, such as a clinically relevant protein or genetic mutation/polymorphism, or quantitatively determine protein or nucleic acid levels in a clinical sample.
All information provided by Luminex Corporation. Luminex and xMAP are registered trademarks of Luminex Corporation.