Patricia F. Fitzpatrick Dimond Ph.D. Technical Editor of Clinical OMICs President of BioInsight Communications

Moving Toward True Portability?

As described by the National Institutes of Health (NIH), point-of-care (POC) instruments combine multiple analytical functions into self-contained, portable devices that can be used by nonspecialists to detect and diagnose disease. They can also enable the selection of optimal therapies through patient screening and monitoring of a patient’s response to a chosen treatment. These devices have attracted public and private funding, as well as considerable investment from large diagnostic companies.

Kalorama Information noted that the emerging area of rapid POC molecular testing attracted over $650 million in investments and financings between 2009 and 2013, alone, while a 2014 Frost and Sullivan report predicted that molecular testing, particularly the overall growth of the global infectious diseases diagnostics market, will approach $12.78 billion in 2018.

The introduction of CLIA waived tests, i.e., assays approved for use by healthcare providers in nontraditional laboratory sites like physician offices, emergency rooms, health department clinics, pharmacy clinics, and other healthcare facilities, is considered a key POC market driver.

Funding that drives the development of POC technologies comes from multiple sources including the NIH, and the U.S. Department of Defense (DOD), as well as private foundations like the Bill and Melinda Gates Foundation and the pharmaceutical industry.

In April of last year, Roche bought POC molecular testing company iQuum for $275 million in cash and the potential for an additional $175 million in milestone payments. The Marlborough, MA company has FDA-cleared and CE-marked diagnostic tests that can be performed at the point of care with minimal training.

While historically focused on infectious diseases that affect developing nations molecular POC has rapidly advanced into other clinical areas, including monitoring of cardiac disease patient drug responses and, most recently, Cepeheid’s GeneXpert (Xpert) test for BCR-ABL, a real-time RT-PCR that can aid in monitoring the p210 BCR-ABL translocation producing results in less than two hours.

Key technologic factors contributing to the favorable funding climate and interest in developing molecular POCs include advances in microfluidics and other instrumentation technologies that can enable development of relatively low-cost platforms and tests that meet the needs of a true POC system.

Specific challenges associated with developing POC molecular tests include the additional steps for sample pretreatment (e.g., cell sorting, isolation, and lysis) and the requirements for nucleic acid extraction and signal amplification, and sample-prep integration with a complete analytical process that does not require skilled operator involvement, and the need for fast results. In particular, true portability remains a moving target, as most current devices tend to be at least shoebox-sized.

First Commercial System

The first commercial version of a real-time nucleic acid detection system was Cepheid’s Xpert system, designed to purify, concentrate, detect, and identify targeted nucleic acid sequences delivering diagnoses from unprocessed samples in approximately 30 minutes. But some scientists have argued that PCR-based devices have fairly large footprints. Andrew St. John and Christopher Price, writing in the August 2014 issue of Clinical Biochemist Reviews, note that existing PCR systems, like Xpert, might, “be seen more as a system to automate PCR-based assays rather than a POC device.”

But, they add, the ability of this relatively small bench-top device to perform real-time quantitative PCR in approximately 90 minutes with minimal operator interaction offers the potential to perform rapid molecular testing in situations where the need for results is urgent.

To date, however, molecular diagnostic POCs might be best characterized as being near the patient rather than at the POC. And while such devices are in development, small, benchtop systems that perform genomic analyses continue to reach the market with an increasing variety of tests that now include infectious diseases, drug responses to drugs used to treat heart disease, and recently, cancer.
Roche’s acquisition of IQuum gave it access to IQuuM’s laboratory-in-a-tube (Liat™) analyzer. It automates all aspects of nucleic acid testing processes, including reagent preparation, target enrichment, inhibitor removal, nucleic acid extraction, amplification, and real-time detection.

The system generates test results in 20 minutes or less to facilitate a treatment decision, according to the company. The analyzer and two initial assays, cobas® Influenza A/B* and cobas® Strep A, are both CE-Marked and FDA-cleared, and the influenza test received a CLIA waiver on May 19.

The Liat analyzer achieves its speed primarily through a proprietary PCR procedure (flow cycling). PCR machines have typically consisted of a large heating block, housing one or more reaction tubes, that cycles its temperature to achieve the target denaturing, annealing, and extension temperatures, a process requiring between 30 minutes and several hours to complete.

The Liat, however, uses a flexible reaction vessel and a modular sample processor, which compresses various segments of the reaction vessel to heat and cool them much faster, allowing results in as little as 20 minutes.

In the cardiovascular space, Spartan Biosciences announced last October that it had received approval from Health Care Canada for its Spartan RX CYP2C19 system, a genetic test that can determine whether patients receiving the antiplatelet medication clopidigrel (Plavix) following percutaneous coronary intervention (PCI) have CYP2C19 mutations that may impair their ability to metabolize the drug. The CYP2C19*2 allele is a common genetic variant associated with increased rates of major adverse events in individuals given Plavix.

The Canadian approval for the test follows the FDA’s 510(k) clearance of the test last August, indicated for use as an aid to clinicians in determining strategies for therapeutics that are metabolized by the cytochrome P450 2C19 gene product, and that are specifically affected by the *2, *3, and *17 alleles. The agency also noted that in the U.S., the device must be used in a clinical lab and is not cleared for POC use.

Spartan says its RX CYP2C19 System is the first near-patient DNA test for personalized medicine that has been approved in Canada. Health Canada also approved its use by healthcare professionals, including doctors, nurses, pharmacists, and laboratory technicians in medical care sites and clinical labs. The device was validated by Spartan and reported in The Lancet. The study enrolled 200 patients in a prospective, randomized, proof-of-concept study using the genetic test to identify carriers of the CYP2C19*2 allele and aimed to assess a pharmacogenetic approach to dual antiplatelet treatment after PCI.

Patients undergoing PCI for acute coronary syndrome or stable angina were randomly assigned to rapid POC genotyping or to standard treatment. Individuals in the rapid genotyping group were screened for the CYP2C19*2 allele. Carriers were given 10 mg prasugrel daily, and non-carriers and patients in the standard treatment group were given 75 mg clopidogrel daily. The authors reported that, based on their results, POC genetic testing after PCI can be done effectively at the bedside and treatment of identified CYP2C19*2 carriers with prasugrel can reduce high on-treatment platelet reactivity.

While the majority of miniaturized systems for nucleic acid analysis use PCR for amplification, newer devices in development may provide truly POC results currently incorporate alternative methods, in particular isothermal amplification methods that do not require thermal cycling.

Isothermal microsystems can be designed for simplicity and low-energy consumption and therefore, proponents say, may outperform PCR in portable, battery-operated detection systems in the future, and in field conditions. These methods include nucleic acid sequence-based amplification (NASBA), loop-mediated isothermal amplification (LAMP), helicase dependent amplification (HDA), rolling circle amplification (RCA), and strand displacement amplification (SDA). 

Moving closer to true portability but still in the development stage, researchers at Michigan State University reported the invention of an inexpensive, user-friendly and compact device, the Gene-Z, an isothermal genetic analysis platform operated by an iPod Touch or Google’s Android tablets. Already validated for microbial analysis in one application, the device could be adapted, its developers say, to detecting cancer in poorer countries.

The system’s features include isothermal amplification of DNA, RNA, and microRNAs, 10–30 minutes assay time, and the potential for GPS and networking of multiple devices. Unique elements of the molecular approach include modifications of microRNAs to be able to amplify and detect via LAMP, use of highly specific genetic signatures extracted from a large number of allelic sequences, use of SYTO 82 dye to track (LAMP using simple photodiodes).

Gene-Z developers say they expect that the device will cost less than $1,000 with disposable chips ranging from $2–$20 based on the density and application.

POC molecular testing technologies garnered more than $650 million in investments and financings between 2009 and 2013. [iStock/BlackJack3D]

Convergent System

Supported by the Defense Advanced Research Project Agency (DARPA’s DxOD project) and the Bill & Melinda Gates Foundation (Grand Challenges), scientists in the departments of physics and bioengineering, University of California Berkeley, developed a microfluidic biomolecular amplification reader (µBAR), an inexpensive, handheld, battery-powered instrument for pathogen genotyping in the developing world. The device, the scientists say, represents a convergence of molecular biology, microfluidics, optics, and electronics technology.

The device reportedly carries out isothermal nucleic acid amplification assays with real-time fluorescence readout at a fraction of the cost of conventional benchtop thermocyclers. The device also features cell phone data connectivity and GPS sample geotagging which can enable epidemiological surveying and remote healthcare delivery.

The µBAR device controls assay temperature through an integrated resistive heater and monitors real-time fluorescence signals from 60 individual reaction chambers using LEDs and phototransistors. Assays are carried out on PDMS disposable microfluidic cartridges which require no external power for sample loading.

As a proof-of-principle, the authors, Frank Myers et al., writing in the August 2013 issue of PLOS One, demonstrated the detection of the HIV-1 integrase gene with the µBAR using LAMP. Since LAMP has previously been demonstrated to work with a range of clinical samples, the scientists say their eventual goal is to develop a microfluidic device which includes on-chip sample preparation from raw samples. The authors note that their goal in developing the instrument was to introduce a platform for isothermal amplification and real-time detection on a multiplexed microfluidic cartridge.

It improves on the state-of-the-art in genetic amplification instrumentation in its cost, operational simplicity, and portability. They point out that their instrument could be conceivably priced ~50× cheaper than conventional real-time thermocyclers. It does not require external power, and at least three assays can be run before the internal battery requires recharging.

In its present form, they concede, the µBAR device does not provide sample-prep capabilities and requires that amplification reagents be mixed off-chip before sample loading. It also requires that DNA be supplied at fairly high concentrations (1,000/uL, for example) because of the small volumes used in the microfluidic chamber. To help further development of POC genetic diagnostic instruments, the authors have provided designs, source code, and instructions for µBAR instruments.

Investigators and clinicians anticipate that given market forces and current demand in specific markets, most notably infectious diseases, portable, hand-held, inexpensive POC devices, equivalent to currently offered full-size molecular analytical systems, will become available. These devices will aid in the detection of mutations or in the identification of infectious agents, as well as aid in the management of patient care.

Patricia Fitzpatrick Dimond, Ph.D. ([email protected]), has been a long-time contributor to GEN and currently serves as the publication’s Technical Editor. She is also president of BioInsight Communications. During her career in the biotechnology industry, she was VP of strategic development and corporate communications at Coley Pharmaceutical Group (now Pfizer), where she developed and managed the company’s investor relations and communications programs.

This article was originally published in the July 2015 issue of Clinical OMICs. For more content like this and details on how to get a free subscription to this digital publication, go to

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