Firefly particles

FirePlex particle technology

Find out how FirePlex® particle technology provides high sensitivity, multiplexed analysis through your flow cytometer.

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FirePlex immunoassays and miRNA assays provide simultaneous measurement of up to 70 proteins or miRNAs at the same time in each well. At the heart of these assays is the innovative FirePlex particle technology, which enables high-performance multiplexing capabilities and easy readout on standard flow cytometers.

The unique architecture of FirePlex particles offers several advantages over standard bead-based assays and other multiplexing technologies:

  • High-plex panels
  • Small sample inputs
  • Broad dynamic range
  • Superior sensitivity
  • Less code bleed, low background and no autofluorescence for higher-quality data from flow cytometry
  • No sample processing or RNA purification required
  • Tolerant to factors that commonly foul other platforms, such as serum albumin, complex glycoproteins, heme compounds, and heparin.


3D analyte binding for a broader dynamic range

Patented FirePlex particles are made from a porous bio-inert hydrogel that provides target capture throughout the 3D volume. Compared with the leading bead-based platform which is limited to 2D surface capture, FirePlex particle technology provides 2x108 more binding sites per particle. More binding sites means assays with a broader dynamic range. For nucleic acid capture and detection, this provides an additional two logs of dynamic range and increased in sensitivity.

With this enhanced performance, more analytes or miRNA can be quantified in the same well, without the need for multiple sample dilutions.


FirePlex particle structure

Particles contain three distinct functional regions each separated from the other by inert spacer regions, thereby minimizing code-bleed to provide superior sensitivity. The central analyte or miRNA quantification region contains probes that capture target proteins or miRNAs. The end regions function as two halves of a barcode to distinguish different particles.

End regions serve as barcodes

  • Emit differing fluorescence intensities to create a fluorescent barcode
  • In a given panel, all particles that capture a particular analyte or miRNA emit the same code

​Central analyte quantification region

  • Contains antibodies or miRNA probes to capture a specific protein analyte or miRNA, respectively​

Particle structure. Distinct regions capture target analytes or miRNA and encode particles (click to expand the image).

How are particles read by the flow cytometer?

Particles passing through a flow cell of a flow cytometer are recorded as three distinct events. The separation between the target quantification region and particle encoding regions reduces signal bleed between channels and eliminates the need for compensation.

FirePlex Analysis Workbench software analyzes flow cytometer data from these three regions to quantify each target analyte or miRNA:

  • Functional regions are triggered in the green channel.

Events from all three regions are detected in the green channel to signify the presence of a particle.

  • Fluorescence intensity in the central region determines protein quantity.

Captured and labeled target analytes or miRNA emit fluorescence proportional to their abundance in the sample. The flow cytometer detects this signal from the center of the particle, through a reporter dye triggering on the red channel.

  • A fluorescent code distinguishes different analytes.

With two end regions of varying fluorescence intensity, a particle code is generated in the yellow channels. Each analyte or miRNA is assigned a unique code, with up to 70 different codes possible. Data from a mixture of particles can be attributed to individual target analytes or miRNA by code identification.

  • Software to stitch all the data together.

After data acquisition, the FirePlex Analysis Workbench software merges events from the three regions of the particles into a single event, returning simply the abundance data for each target analyte or miRNA.

>> Download the FirePlex Analysis Workbench software for immunoassays

>> Download the FirePlex Analysis Workbench software for miRNA assays

True multiplexing ability for comprehensive biomarker profiling

With the ability to assign up to 70 different codes, FirePlex particles can distinguish significantly more analytes or miRNA within a single panel than conventional bead-based immunoassays for flow cytometers.

A particle mixture is added to each well, containing on average 20 particles specific for each analyte or miRNA, with up to 70 different targets possible. This generates 20 reads for every analyte or miRNA, with a maximum of 1,400 data points per well—providing reliable, high-quality results.

True multiplexing. Dozens of protein analytes or miRNAs are detected and quantified within a single well.

The unique properties of FirePlex particles enable you to measure exactly the analytes and miRNAs you need with excellent performance across a breadth of sample sources. Our FirePlex miRNA and immunoassays profile biomarkers directly from very small volumes of crude biofluids—requiring only 12.5 µL for immunoassays or 10 µL for miRNA assays.  With FirePlex, you get true multiplexing for both proteins and miRNAs without compromising performance.

Learn more about our FirePlex assays


Compatible flow cytometers

FirePlex immunoassays can be conveniently run on your own existing flow cytometer, eliminating the need to purchase dedicated equipment. FirePlex particles are validated for use with the following flow cytometers fitted with green, yellow and red detectors.

  • Millipore's Guava EasyCyte™
  • BD Biosciences' Accuri™ C6
  • ThermoFisher's Attune® (limited analyte list; not validated for the NxT)
  • BD Biosciences' FACSCanto™, LSRII, and LSRFortessa™
  • Check with our team whether your flow cytometer is compatible


>> View our full list fo validated flow cytometers


Our Cytometer Setup kit (ab245835) can be used to ensure your cytometer is compatible and has appropriate settings.

 
Don't have access to a compatible flow cytometer? You can still profile cytokines with the help of our sample profiling team.


FirePlex® is a registered trademark in the United States and is an unregistered trademark elsewhere.




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