Frequently Asked Questions Mass Spectrometry Services

  • What is the xMAP® technology?

The Multiplex Analyte Profiling (xMAP®) technology was developed by the Luminex® Corporation around proven flow cytometry, microspheres and traditional chemistry technologies. This technology features a flexible, open-architecture design and allows performance of a wide variety of bioassays accurately, quickly and cost-effectively. The technology allows simultaneous analysis (multiplexing) of up to 100 analytes in a single sample well through the combination of fluidics, optics, lasers, temperature control, software, high-speed digital processors and xMAP microspheres (beads). These 100 distinctly fluorescently dyed 5.6 micron-sized polystyrene beads serve as identifiers and a solid surface which enables performing a fast and cost-effective bioassay using relatively small sample volumes. Through the use of two lasers, the first red laser exciting the dye inside the bead and the second green laser exciting the fluorophore bound to the surface of the bead, the photo diode detectors can measure the excitation emission intensities inside the beads and a photomultiplier tube detects the excitation emission intensities on the surface of the bead. These intensity measurements are analyzed through high-speed digital processors and advanced computer algorithms, which allow for real-time analysis and quantification. This xMAP® platform can be used for quantitative analysis of DNA, RNA and protein in biological and clinical samples.

  • What are the xMAP® microspheres (beads)?
    • The polystyrene beads are internally dyed with red and infrared fluorophores. In order to create 100 different microspheres, different quantities or ratios of the red and infrared dyes are implanted into the beads, giving each bead a unique spectral signature. Recently, magnetic beads have also been introduced. The magnetic beads called MagPlex Microspheres® by Luminex™ are super-paramagnetic microspheres that are internally labeled with fluorescent dyes and contain surface Carboxyl groups for covalent attachment of ligands (or biomolecules).

What are the benefits to xMAP® technology
Time and money savings through:
a) Multiplexing – Run up to 100 bioassays simultaneously in one sample well allowing for parallel analysis and efficient utilization of small amounts of samples.
b) Customized assays – Allow each scientist to develop assays for their individual needs.
c) Flexibility - Small sample amounts and multiple sample types allow assays to be tailored to the scientists’ requirements.
d) Cost/target – lower cost per target when compared to cost of performing PCR and ELISA for multiple targets.

How does the Panomics QuantiGene® Plex 2.0 assay for RNA work?
This assay combines branched DNA (bDNA) signal amplification technology, which amplifies the reporter signal instead of the target sequence, and xMAP® microspheres (beads) to provide accurate RNA profiling. First, RNA is released from the cells through cell lyses. Samples are then hybridized overnight, where mRNA transcripts are captured to the appropriate beads. After hybridization with a pre-amplifier and amplifier, Streptavidin-conjugated Phycoerythrin (SAPE) is bound to the mRNA transcripts bound to the bead. The lasers in the Luminex 200™ instrument measure the SAPE fluorescence, which is proportional to the amount of mRNA transcripts.

  • What kind of sample types are compatible with the Panomics QuantiGene® Plex 2.0 assay?

Cultured cells, whole blood, blood collected in PAXgene tubes, dried blood spots, fresh/frozen tissues, FFPE samples, and purified RNA can be analyzed.

  • How many RNA targets can be analyzed in each well for the Panomics QuantiGene® Plex 2.0 assay?

At this time up to 36 targets can be quantified in each well 

  •  Is it necessary to isolate or purify RNA prior to performing the Panomics QuantiGene® Plex 2.0 assay?

No, there is no need for RNA isolation/purification, reverse transcription, or amplification by polymerase chain reaction (PCR).

  • How long does a gene expression profiling service take?

Turnaround time depends on what is required and number of samples. Please call usfor more details.

How sensitive is the Panomics QuantiGene® Plex 2.0 assay?
This assay has a Limit of Detection of ≤ 1,000-2,000 transcripts/assay well, a Limit of Quantitation of ≤ 2,000-4,000 transcripts/assay well and a Linear Dynamic Range of ≥ 3.5 logs.

Are housekeeping genes required for the Panomics QuantiGene® Plex 2.0 assay?
Housekeeping genes are not required. Although not required, it is recommended to include housekeeping genes, or control genes, since this allows for normalization.

How many RNA samples can be run on the Panomics QuantiGene® Plex 2.0 assay?
The Panomics QuantiGene® Plex 2.0 assay for RNA uses a 96 well plate, with 3 wells used for assay standards and background controls; therefore, 31 samples can be run in triplicates. More samples can be analyzed if single or duplicate assays are carried out.  

  • How many protein samples can be run on multiplex assay (xMAP®)?

The multiplex protein assay uses a 96 well plate, with 9 wells used for assay standards and background controls; therefore, 29 samples can be run in triplicates. More samples can be analyzed if single or duplicate assays are carried out.

What kind of assays currently exist? ?
Multiplex assays are available for human, mouse, rat and monkeys. There are assays targeting multiple mechanisms and pathways. Off-the-shelf assays are available to quantify the following:

  • Cytokines
  • Angiogenic factors
  • Growth factors
  • Inflammatory and Acute phase proteins
  • Apoptotic proteins
  • Kinases
  • Neurobiology proteins
  • Cancer biomarkers.

 

What types of samples are compatible with the multiplex assay (xMAP®)?
This assay can use the following sample types: tissue homogenate, whole blood, cell lysate, serum and  plasma. Purified proteins and RNA can also be analyzed.  

What is the cost of an xMAP® or QuantiGene® assay?
The cost for an assay depends on the type of target, number of targets and the number of samples that will be analyzed. If we know what you want to measure, we can provide you with a competitive quotation within 24 hours.