XRpro® Technology

Label-free Platform for Direct Ion Flux Measurement

Overview

XRpro® technology leverages the unique capabilities of x-ray fluorescence for high throughput analysis of plasma membrane ion channels and transporters, including nonelectrogenic symporters and antiporters. XRpro® technology is a label-free technology that directly quantifies ion flux in cell populations without dyes, fluorophores, and radiolabels and offers the ability to conduct assays in complex buffers and media, including 100% serum. All elements with an atomic number of 13 (aluminum) or greater are measured simultaneously, including biologically important monovalent ions (e.g. K+), divalent ions (e.g. Ca2+), transition metals (e.g. Zn2+) halogens (e.g. Cl-), and tracer ions (e.g., Rb+, Sr2+).

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Technology Time

XRpro® Label-free Technology

XRpro® technology is a novel application of X-ray fluorescence that addresses many of the challenges of conventional ion channel and transporter assay technologies through direct measurement of elements.

XRpro® Technology Applications

XRpro® technology leverages the unique characteristics of X-ray fluorescence for drug discovery. In a few seconds or less, XRpro® instruments measure an entire X-ray fluorescence spectrum for each sample and measure all elements with an atomic number greater than 12 (Al and higher). Elements shown in blue can be directly measured, opening the door for a wide range of biochemical analyses. Target applications include ion flux assays for monovalent and divalent cations, halogens, first-row transition metals, and small molecules. No large organic fluorophores or radioactivity is required. 

The ability of XRpro® technology to simultaneously analyze multiple elements enables two experimental approaches – direct measurement of target ions and the use of tracer ions. Tracer ions are chemically similar to biological ions and are transported by the same channels and transporters, but have distinct element content. Use of tracer ions eliminates background, and allows specific measurements of ion flux that takes place during experiments. For example, all living cells contain a background of K+ and Ca2+. Using Rb+ and Sr2+ as tracers for monovalent and divalent ion flux increases signal to noise and simplifies cell biology by eliminating the need to try to reduce K+ and Ca2+ background. 

Technology Benefits

The strengths of XRpro® technology are derived from the physical properties of X-rays and the large number of elements that can simultaneously be analyzed. XRpro® enables direct measurement of ions and metals that are otherwise difficult to address. This benefit opens the door to analysis of nonelectrogenic systems, transporters, and other targets that were not previously possible or cost effective on a large scale, including a large number of inorganic transporters.

The penetrating ability of X-rays and the lack of dyes make XRpro® insensitive to the chemical environment of target ions. It is compatible with complex buffers and media, including 100% serum, that are challenging for other technologies. This feature enables high throughput, direct activity-based measurements to identify reductions in compound efficacy due to serum protein binding.

X-ray fluorescence, a label-free technology analogous to optical fluorescence that biochemists use every day.  The fundamental difference is the use of X-ray photons, rather than optical or ultraviolet, for excitation.  The result is that individual atoms fluoresce, without the need for organic fluorophores.  Each element on the periodic table generates a unique set of X-ray emission lines that can identify and quantify the atoms present in a sample.

X-ray fluorescence is generated when X-ray photons from the excitation source have enough energy to eject inner-shell electrons (Figure 1) from atoms in the sample.  Electrons from higher orbitals (Figure 2) transition to fill the gaps left by the ejected electrons, emitting X-ray photons in the process.  Because the energy differences between orbitals are dependent on the atomic number of the element (Z), each element emits X-rays with different, characteristic photon energies.  In addition, because only inner-shell electrons are involved, and because X-rays penetrate matter, X-ray fluorescence signals are not altered by chelation, binding, oxidation state, or complex chemical mixtures.

In an XRpro® instrument, an energy resolving, photon counting X-ray detector is used to measure the intensity of X-ray fluorescence.  A full spectrum (Figure 3) is measured for each sample, allowing simultaneous measurements of multiple elements.  Count rates are linear with concentrations over more than four orders of magnitude. 


  • Cell based Assays

  • Cell based Assays

XRpro® technology leverages the unique characteristics of X-ray fluorescence for drug discovery.  In a few seconds or less, XRpro® instruments measure an entire X-ray fluorescence spectrum for each sample and measure all elements with an atomic number greater than 12 (Al and higher).   Elements shown in blue can be directly measured, opening the door for a wide range of biochemical analyses.  Target applications include ion flux assays for monovalent and divalent cations, halogens, first-row transition metals, and small molecules.  No large organic fluorophores or radioactivity are required.

  XR Pro App

XR Pro AppThe ability of XRpro® to simultaneously analyze multiple elements enables two experimental approaches: Direct measurement of target ions, and the use of tracer ions. Tracer ions are chemically similar to biological ions and are transported by the same channels and transporters, but have distinct element content. Use of tracer ions eliminates background, and allows specific measurements of ion flux that takes place during experiments.  For example, all living cells contain a background of K+ and Ca2+.  Using Rb+ and Sr2+ as tracers for monovalent and divalent ion flux increases signal to noise and simplifies cell biology by eliminating the need to try to reduce K+ and Ca2+ background.

 

ChartThe strengths of XRpro technology are derived from the physical properties of X-rays and the large number of elements that can simultaneously be analyzed.  XRpro enables direct measurement of ions and metals that are otherwise difficult to address.  This benefit opens the door to analysis of nonelectrogenic systems, transporters, and other targets that were not previously possible or cost effective on a large scale, including a large number of inorganic transporters.

The penetrating ability of X-rays and the lack of dyes make XRpro insensitive to the chemical environment of target ions.  Itis compatible with complex buffers and media, including 100% serum, that are challenging for other technologies.   This feature enables high throughput, direct activity-based measurements to identify reductions in compound efficacy due to serum protein binding.

XR pro Tech Benefits

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