Existing and Emerging Technologies for Point-of-Care Testing

Abstract

The volume of point-of-care testing (PoCT) has steadily increased over the 40 or so years since its widespread introduction. That growth is likely to continue, driven by changes in healthcare delivery which are aimed at delivering less costly care closer to the patient’s home. In the developing world there is the challenge of more effective care for infectious diseases and PoCT may play a much greater role here in the future. PoCT technologies can be split into two categories, but in both, testing is generally performed by technologies first devised more than two decades ago. These technologies have undoubtedly been refined and improved to deliver easier-to-use devices with incremental improvements in analytical performance. Of the two major categories the first is small handheld devices, providing qualitative or quantitative determination of an increasing range of analytes. The dominant technologies here are glucose biosensor strips and lateral flow strips using immobilised antibodies to determine a range of parameters including cardiac markers and infectious pathogens. The second category of devices are larger, often bench-top devices which are essentially laboratory instruments which have been reduced in both size and complexity. These include critical care analysers and, more recently, small haematology and immunology analysers. New emerging devices include those that are utilising molecular techniques such as PCR to provide infectious disease testing in a sufficiently small device to be used at the point of care. This area is likely to grow with many devices being developed and likely to reach the commercial market in the next few years.

Introduction

The dominant model of laboratory testing throughout the world remains the centralised laboratory in which more and more of the analytical processes are automated to enable the analysis of large numbers of samples at relatively low cost. This trend is well established in biochemistry and haematology and is now extending to other disciplines including microbiology and anatomical pathology.

However healthcare is changing, partly as a result of economic pressures, and also because of the general recognition that care needs to be less fragmented and more patient-centred.¹ Many countries are facing the reality of having to limit the growth in healthcare budgets or in some cases reduce healthcare spending. One way to achieve this goal is to reduce relatively expensive care in secondary and tertiary hospitals and encourage more patients to be assessed and treated in primary care or the community. It is uncertain whether the central laboratory concept is best suited to the needs of this more primary care orientated care model which is still in its infancy. Alternative models using PoCT are being increasingly considered, particularly for people in more remote locations such as found in Australia,² but also in relatively densely populated countries such as the UK where telehealth applications are being actively considered to provide more care in the patient’s home.³

The need to make healthcare more patient-centred is also a global trend and is based on the premise that healthcare should be organised more around the patient rather than the provider. Centralised testing does not represent a convenient process for many patients with the testing process often being disconnected from the consultation process such that more than one visit to the doctor is required to complete the assessment process. This problem applies particularly to those with a chronic disease such as diabetes who require regular monitoring including frequent blood tests. The growth in self-monitoring of blood glucose, which is by far the largest segment of PoCT, is in part a testament to this need for more convenient and, in some cases, more effective care.⁴

Pressure on healthcare budgets and the trend to patient-centred care might be perceived as problems confined to the developed world. While true in part, the pace of development in countries such as India and China means that they have a growing middle class with its attendant healthcare problems of chronic disease such as diabetes and cancer. This will lead to the same problems as those being faced by westernised healthcare systems. But poverty and disease remain significant problems in the developing world with many infectious diseases leading to significant mortality. Effective diagnostic testing has been difficult to achieve in this area but PoCT is seen as one way to meet this need and several major global initiatives are devoted to providing such testing.^5,6 Consequently it is likely that PoCT will increase substantially in developing countries in the next decade.

Market Growth of Point-of-Care Testing

Various reports are available to document the growth in in vitro diagnostics (IVD) markets including various categories such as PoCT. While the numbers vary between reports, the total IVD market was believed to be worth US$51 billion in 2011 of which approximately US$15 billion was PoCT. The latter is projected to show compound annual growth of 4% to reach US$18 billion by 2016. Of the total PoCT market in 2011, 55% of it was in the US, 30% in Europe and 12% in Asia.⁷

The PoCT market is made up of what is termed ‘over the counter’ products such as glucose monitoring and pregnancy testing (also called non-professional testing) and the ‘professional market’ which includes all other testing including critical care, infectious disease, cardiac markers, diabetes, lipids, coagulation and haematology. In 2011 the professional market was worth US$5.66 billion and is projected to grow to US$6.76 billion by 2016. Glucose testing (by healthcare providers rather than patients) is the largest sector followed by pregnancy and critical care testing while infectious disease testing is the fastest growing area.⁷

Data on the growth of molecular testing is also interesting in the context of the increase in infectious disease PoCT.⁸ The total molecular diagnostics market is believed to be worth US$4 billion in 2011 and grow to US$7 billion by 2016 with compounded annual growth rates in infectious disease testing of more than 18%. With the emergence of new devices to perform molecular testing at the point of care and the well-documented needs for more infectious disease testing in the developing world, it will be interesting to see how much of that growth takes place in point of care locations rather than the central laboratory.

Required Features of PoCT Devices

Designers of PoCT devices start with the needs of their users and these needs will to some extent depend on the clinical setting. However some features are common to virtually all users in all settings. As documented by St John et al.,⁹ these key requirements include:

1. Simple to use.

2. Reagents and consumables are robust in storage and usage.

3. Results should be concordant with an established laboratory method.

4. Device together with associated reagents and consumables are safe to use.

With the growing potential for PoCT to improve healthcare in the developing world, particularly through timely detection of infectious diseases, developers of such devices have been guided by more specific design criteria. These are to ensure that the technology can address the needs of the user in a clinically and cost effective manner and avoid the introduction of possibly expensive devices which fail to deliver the required outcomes. Thus the World Health Organisation (WHO) has provided guidelines for those developing PoCT devices for the detection of sexually transmitted infections (STI), a major health problem in the developing world, and in the developed world for diseases such as Chlamydia and HIV.¹⁰ These guidelines are known as ASSURED and are shown in Table 1.

Table 1.

The ASSURED guidelines that indicate the features that should be designed into all PoCT devices.

Affordable – for those at risk of infection Sensitive – minimal false negatives Specific – minimal false positives User-friendly – minimal steps to carry out test Rapid & Robust – short turnaround time and no need for refrigerated storage Equipment-free – no complex equipment Delivered – to end users

The reality of course is that it has been difficult to deliver on all of these technology requirements and a recent paper discusses what compromises and trade-offs would be acceptable to developed world healthcare providers working in the areas of STIs. Hsieh et al. conducted an online survey following a focus group involving clinicians and others offering STI services, to discuss what would be the ideal PoCT device for this area.¹¹ The survey included a section entitled ‘build your own PoCT’ using a discrete choice experiment approach, a tool from marketing research that allowed participants to identify which of the ASSURED features they would chose over others. The results of this analysis identified that a longer time to getting a result from the device or TAT was a major barrier to greater use of PoCT for STIs such as Chlamydia and HIV. Irrespective of the test organism being detected, sensitivity of 90–99% was the most important attribute followed by a low cost of US$20 and short detection time of 5 min. However another survey conducted by the same group indicated that industry professionals preferred a 15 min turnaround time. Huang et al. in a comparative effectiveness study found that women attending an STI clinic would be willing to wait for up to 40 min for their result.¹² For detection of early HIV and syphilis, sensitivity was still the most important but specificity was ranked second.¹¹ Given the compromises inherent in virtually all measurement technologies, it may be useful to extend needs assessments like these to other areas of PoCT.

For PoCT devices that will be used in the developed world, some of the ASSURED criteria will also remain relevant but others will be substantially different. Thus instead of being equipment free, the need will be for relatively sophisticated equipment that at a minimum can provide a quantitative result, presentation of the results, decision support and, ideally, connectivity to other information systems such as the patient’s electronic health record. While the technology to provide all these features undoubtedly exists, they come at a cost which may be difficult to recover using the most common business model for PoCT which is that used for the central laboratory, based on complexity and reagent costs, thus only charging for the test strips/cartridges.¹³ When one combines these equipment needs with what are seen as other competitive requirements such as a small sample volume, whole blood, production of a result within 10 min of applying the sample, ease of use and requisite analytical performance, it is possible to appreciate the technological challenges involved in building such devices.

In recent times a newer challenge has arisen, namely the ability to simultaneously measure multiple analytes on the same cartridge, or multiplexing as it is known. Multiplexing is a rather broad and undefined term but, with the exception of devices used in critical care such as blood gas analysers, the number of multi-analyte PoCT devices is relatively few. Of the few that have appeared, those that have the ability for example to measure multiple cardiac enzymes or several different type of drugs are not universally popular with users because they will be charged for all such parameters, irrespective of whether they need the complete panel. However as more healthcare moves away from the hospital into the community, the demand grows for multi-analyte point of care platforms since these avoid the need for several devices, all with the attendant needs of multiple training, quality management and interfacing processes and the increased risk of errors. But the technological challenges of meeting all these needs has meant that the introduction of new technologies has been relatively slow with the largest sections of the current PoCT market such as glucose, international normalised ratio (INR), cardiac markers and blood gases all using technology that was introduced at least 20 years ago and sometimes much longer.¹³

Types of PoCT Technology

A typical classification of PoCT technology splits devices into small handheld ones including quantitative and qualitative strips, and those which are larger bench-top devices with more complex built-in fluidics, often variants of ones used in conventional laboratories. It is possible to identify a number of key design components that are incorporated into all devices, the collective aim of which is to achieve, as far as possible, all the desired features that were discussed in the previous section.⁹ Although not all are present in the simpler devices such as dipsticks, these key design components are shown in Table 2. With the trend of increasing miniaturisation of devices and the application of technologies developed in relation to consumer electronics, it is becoming increasingly possible to make smaller and smaller devices that incorporate all of these key design features.

Table 2.

Key design components of PoCT devices.

Operator interface Bar code identification system Sample delivery devices Reagent storage and availability Reaction cell Sensors to detect the measurement reaction Control and communication systems Data management and storage Manufacturing requirements

Several comprehensive reviews of PoCT technologies exist in the literature, and these remain reasonably current due to the relatively slow pace of new technology development and slow adoption by users.^9,14 The commercially available forms of these two major types of devices are briefly discussed below, highlighting some of the latest versions of these technologies. This is followed by a review of technologies which are in development and show promise but are not generally yet available for routine use.

Small Handheld PoCT Devices

A myriad of small devices exist for PoCT that range from the humble, so-called dipstick to the sophisticated, small cartridge devices used in blood gas analysis. These devices are truly portable and typically used by the patient themselves or by healthcare professionals at locations adjacent to the patient such as by the bedside, in the clinic or in the patient’s home. Their portability means that how these devices are used, sometimes referred to as the operational workflow, is usually different to the use of larger or bench-top devices. For example, small, portable devices often use fingerstick, capillary samples which are applied directly to the PoCT instrument without the need for sample containers, labelling or transport which are required for analysis by a larger bench-top PoCT instrument some distance from the patient. While avoiding these additional steps is highly convenient, there are risks associated with such procedures and the testing process must be designed to minimise such risks with appropriate training and documentation.

The dipstick is a PoCT technology that has stood the test of time and is used frequently today by patients, nurses and doctors in many different locations. In its simplest form a urine sample is applied to a porous pad containing reagent and reflectance technology is used to provide a semi-quantitative estimate of the analyte.¹⁵ Dipsticks can detect one or up to 10 analytes and can be used in conjunction with a small reading device in order to reduce potential operator error. To measure analytes in blood the pads contain several layers, one of which is a membrane that prevents red cells from entering the part of the pad where reflectance is measured.¹⁶ Dipsticks of this type require more operator care because their performance is dependent upon sample volume as well as the need to cover all the pad, and reading the result after a certain time has elapsed after applying the sample – although even in the case of the latter point, the issue of reading time has been overcome. Such devices can measure a clinically useful range of analytes in urine and whole blood.¹⁷

Immunostrips are immunosensors where the recognition agent is an antibody that binds to the analyte with detection by reflectance or fluorescence spectrophotometry. They have been built in a variety of different formats, the first of which was a flow-through design using a porous matrix cell in which a heterogeneous immunoassay enabled the measurement of β-hCG at the point of care.¹⁸ Lateral flow designs where the separation takes place as the sample moves along a solid phase are much more common, and in fact are the dominant technology in this sector of the PoCT market. The principles behind lateral flow devices are well described in the literature and over the years, with increasing knowledge about solid phase and surface chemistry technology, the capability and reliability of these devices has improved considerably.¹⁹ These days they also have built in quality control checks which indicate whether the strip technology is working correctly. The utility of lateral flow strips can be extended from qualitative to quantitative measurement through the use of small reader devices that incorporate multichannel light detectors. The latter are often a charge-coupled device (CCD) or CCD camera which can measure much lower light signals than a conventional reflectometer. These devices are commonly used to measure cardiac markers and other acute care parameters such as D-dimer.

The continuing development of strip technology together with meter-type readers is best exemplified by the ubiquitous glucose meter which is by far the largest portion of the PoCT market. Current models reflect several decades of innovation since their introduction in the 1970s, both in terms of strip technology and also in meter design. The overall goals have been to make them easier to use, with less potential for errors, and to minimise the effects of interferences.²⁰ Of the many different types of strip that have been developed, all are biosensors incorporating an enzyme such as glucose oxidase, glucose dehydrogenase (GDH) or hexokinase. They are often termed thick-film sensors because each strip is composed of several layers each with designated functions such as separation, spreading or support.

There are two types of detection systems: photometric or electrochemical. The latter detection systems have enabled the design of strips that are less subject to interference although problems still persist. Strips that use glucose oxidase are more substrate-specific but are affected by oxygen tension, with high PO₂ values leading to falsely low results. Blood oxygen tension does not affect GDH-based strips but GDH strips that use the pyrroloquinolinequinone form of the enzyme are subject to interference by maltose. Excessive concentration of the latter such as in some parenteral nutrition solutions can cause falsely high glucose results when the patient is actually hypoglycaemic, and several deaths have been attributed to these inaccurate results.²¹ Haematocrit is another important interference, although the effects have been reduced by some newer strip designs. Many different factors can lead to inaccurate glucose results from strip tests and a comprehensive review of these is provided by Tonyushkina et al.²²

Electrochemical detection of glucose also made possible more compact meter designs, smaller sample requirement, the production of non-wipe strips and more rapid result delivery – which are all features that facilitate an easier measurement process, particularly important for patients who are self-monitoring. A major cause of inaccuracy in many meter-based systems is failure to insert the correct calibration code into the meter for a particular batch of strips, and this can cause errors of up to 30%. Now some strip and meter designs incorporate automatic coding and calibration processes.²³

A number of guidelines have been published which document the required accuracy of glucose meters, namely the level of agreement between meter results and those from conventional laboratory testing (Table 3). The most commonly cited guideline is from the International Organisation for Standardisation (ISO) 15197:2003, which states that 95% of results <4.2 mmol/L by meter should be ±15% of the laboratory method and for results ≥4.2 mmol/L the difference should be ±20%.²⁴ There have been calls to tighten these accuracy standards, primarily for meters used in hospitals where insulin regimens are generally based on meter results. A modelling study by Boyd et al. showed that using the meters available at that time would lead to a significant number of errors in dosing.²⁵ Thus a recent Clinical and Laboratory Standards Institute (CLSI) guideline, POCT12-A3, indicates a requirement for meter results to be within 12.5% of laboratory results²⁶ while the US Food and Drug Administration (FDA) has stipulated a requirement for agreement between meters and laboratory results to be ±10.0%.²⁷ An evaluation of the newest designs from all the major manufacturers showed that only one current meter, the Optium Xceed (Abbott Diabetes Care), would meet the criteria of ±12.5% for results <4.2 mmol/L (Table 3).²⁸ No doubt innovation and incremental improvements in technology will lead to newer strip and meter designs which enable their performance to be improved still further.

Table 3.

Recommended guidelines for the accuracy of glucose meters compared to laboratory results and the accuracy of results in a recent evaluation of six different glucose meters (ref. ²⁸).

Comparison of glucose meter performance stipulated in various guidelines
Guideline	Glucose range (mmol/L)	Performance criteria – meter compared to laboratory results
ISO 15197 - 2003	<4.2	95% results ± 15%
ISO 15197 - 2003	≥4.2	95% results ± 20%
CLSI POCT12-A3	>5.5	95% results ± 12.5%
FDA recommendation	>3.8	99% results ± 10%
Glucose meter performance in a recently published evaluation
Meter	% results within 20% at 4.2 mmol/L	% results within 12.5% at 5.6 mmol/L
Optium Xceed	100%	98%
Accu-Chek Performa	98%	86%
Contour TS	98%	94%
Nova Max	93%	83%
Nova StatStrip	99%	93%
OneTouch Ultra 2	97%	86%

Another group of common meter-type devices used by patients and healthcare providers are ones that measure prothrombin time reported as INR levels as part of monitoring warfarin therapy. First devised some two decades ago, a number of different technologies have been developed to measure INR at the point of care including optical and electrochemical detection. Innovation has been applied both to the strip, where a drop of blood is placed, and to the meter into which the strip is placed. All of these innovations have improved the reliability of these devices to the extent that a systematic review of the literature on the quality assessment of devices for self-monitoring indicated that they generally provide comparable results to laboratory-generated INR values.²⁹ However it is important to emphasise this review only looked at studies of self-monitoring and not at PoCT INR meters used by healthcare professionals where the range of expected results may be larger than that of self-monitoring and a different level of performance may be required.

So-called integrated cartridges are an important sub-class of handheld devices and best exemplified by the i-STAT device. After placement of a small sample of whole blood the cartridge is inserted into a reader for measurement. The cartridge utilises thin-film sensors in combination with microfluidics and cartridges are produced in various formats for different analytes.³⁰ Now more than two decades since their introduction, the devices are used in many different point of care locations. Their popularity is due to the extensive critical care testing menu that is available on a single device, albeit using a range of different cartridges. Thus users need to be familiar with only one operating procedure rather than multiple ones when using several devices. Thus the i-STAT represents an economical way to provide relatively low numbers of critical tests.

The Epoc critical care testing system is also a handheld critical care testing analyser but of a very different construction being based on so-called Smart Card Technology.⁹ Here the biosensors and microfluidics are printed on a 35 mm tape-on-reel format which does not demand the clean air type of manufacturing that is required for thin-film sensors. Once again, a number of different cartridges are available to provide a range of critical care tests with the advantage of operators having to be familiar with the use of only a single device.

Larger Bench-Top PoCT Devices

The design of this type of instrument overlaps with some of those that are used in the central laboratory. Laboratory devices first migrated periodically to outpatient clinics for services such as diabetes care. As more of this care is undertaken in locations outside of the hospital, these devices are now being used in a wide variety of point of care locations. However space is often at a premium in such locations so devices have to be reduced in both size and complexity so that they can be operated by non-laboratory trained staff. The general trend of miniaturisation and increasing computer processing power is facilitating the development of solutions to meet these needs.

It is important for all PoCT devices to be sufficiently accurate and precise to meet the clinical purposes for which they are designed. In the case of HbA_1c measurement it has been difficult to produce a PoCT device that is small, simple to operate and meets the analytical goals. Thus small handheld devices for HbA_1c measurement have been wanting in terms of both accuracy and precision.^31,32 As a result larger, bench-top analysers are required to meet the required analytical specifications, but successive product developments in this area show increasing compactness and capability. The first of this type of instrument was the Siemens DCA which has the capability of measuring both blood HbA_1c and urine albumin/creatinine ratio.³³ More recently the Alere Affinion device became available, of similar size, with the same tests as the DCA but with the addition of C-reactive protein (CRP).³⁴ The most recent instrument of this type is the Roche cobas b101 which can measure HbA_1c and lipids and is significantly smaller than the DCA and the Affinion. What all three devices share is a very simple user interface that easily allows a small droplet of whole blood to be added to a small reagent cartridge or disc that contain the reagents. This is then slotted into the instrument and the analysis proceeds without further involvement of the operator. For all instruments, each test requires a separate cartridge or disc. In the case of the cobas both the lipid and HbA_1c tests can be run sequentially within 15 min, each test taking approximately 6 min. Several evaluations have shown that all three instruments meet the required analytical performance for both HbA_1c and lipid monitoring.^33–35

Small desktop instruments are also available to measure a wider range of general chemistry analytes such as the Piccolo which uses a small disposable rotor that contains all the required reagents and diluents to perform a particular or a related group of tests such as liver function tests.³⁶ The ability to measure a group of tests together confers a degree of uniqueness to the Piccolo but possibly because of expense, this type of instrument has had limited success outside of the US Doctor’s Office market. However there is still a perceived need for devices that can run a wider range of analytes such as liver function tests as well as the diabetes-type tests and certain immunoassays on a single sample/ cartridge or cassette. A newer entrant into this market is from the Samsung Corporation which advertises a number of different PoCT analysers called LABGEO and one of these is available in Australia. The LABGEO PT10 has the capability of measuring up to 15 chemistry parameters using one reagent cartridge while other cartridges offer a smaller combination of analytes. However there is only limited information available in the public domain about the measurement technology or about its analytical performance.

Samsung also offer PoC instruments for immunoassay measurement such as the LABGEO IB10 which can determine cardiac markers and this has been evaluated for use in the Scottish ambulance service.³⁷ The measurement technology is based on lateral flow technology in the form of a compact disc and developed by Nanogen through its Nexus-DX operation.³⁸ Nanogen claim to have made a number of enhancements to the lateral flow process including the use of europium as the signalling agent and collectively these enhancements contribute to improved assay performance. As yet no peer-reviewed publications are available to demonstrate the performance of this newer PoC immunoassay device.

Blood gas analysers remain the dominant application in the bench-top portion of the PoCT market. Nearly 50 years of innovation have produced considerable change in the design and capability of these devices which, because of their extended menu, are now more correctly termed critical care analysers. The menu of these devices can now include electrolytes, urea, creatinine, glucose, lactate and bilirubin as well as haemoglobin derivatives but the other major change has been the move to cartridge-based technology. The major difference here to the cartridge-based systems used in hand-held devices is of course that the sensors are designed to be reusable. Constructed using thick-film technology, the sensors for each of the tests and all the required reagents for calibration and washing are packaged into a single cartridge pack which is inserted into the instrument.⁹ The lifespan of the cartridge when inserted into the machine is based on the number of samples analysed or an elapsed period of time. The manufacture of such complex combined sensor and reagent systems with sufficiently extended shelf-lives to be economical was quite challenging and periodic product recalls are in part a reflection of those challenges. But these types of cartridge-based systems are now common throughout the world, showing that routine and reliable manufacturing of these systems is now possible. The fact that they are widely used despite being more expensive than older generation systems is a reflection of the desire for convenience that comes from such low maintenance systems. Table 4 lists some of the features which are now common in many of these critical care systems and which facilitate their use by non-laboratory trained personnel.

Table 4.

Features of critical care analysers that contribute to ease-of-use and reduce the risk of errors.

Long-life, maintenance-free electrodes or disposable sensor packs Touch screens as the user interface Software that can demand user and patient identification Built-in bar code scanners Sample aspiration instead of injection Reduced sample sizes Clot detection within analysis chamber Sample detection to prevent short samples Liquid calibration systems instead of gas bottles Automated calibrations Automated quality control sampling Sophisticated QC programs including interpretation of data Connectivity to information systems allowing remote monitoring and control Built-in videos for training purposes

Other time critical parameters which may need to be determined at point of care are analytes such as troponin, b-type natriuretic peptide, D-dimer, β-hCG and CRP. Some instruments dedicated to measuring these parameters utilise lateral flow strips in conjunction with a reader, as discussed previously. A different and unique approach is the Radiometer AQT90, a bench-top, random access immunoassay analyser utilising a cup containing dried reagents incorporating antibodies to the analytes of interest and europium lanthanide chelate as the signal reagent.³⁹ Sample is added via a unique, walk-away, closed-tube sampling device and time-resolved fluorescence is used to detect the analyte.

While the menu of point-of-care tests continues to be primarily clinical chemistry parameters, PoCT devices are also becoming available for haematology and immunology tests. Haematology instruments range from ones that measure just haemoglobin and white cell counts to the Sysmex pocH-100i which can measure up to 17 parameters but is much reduced in size compared to its laboratory equivalent.⁴⁰ The QBC Star instrument is an example of a so-called dry haematology system, using dry reagents as opposed to bulky wet reagents thus lending its use to PoCT locations. Samples are placed into a special QBC Star haematocrit tube system which is placed into the desk-top instrument that incorporates a centrifuge, a dual light source and an optical detector system. The instrument produces a 9-parameter blood count and an evaluation showed comparable measurements to that of a conventional laboratory analyser.⁴¹ While the evaluation was performed by laboratory staff, their recommendation was that it is suitable for use in a wide variety of locations outside of the laboratory.

Another relatively new entrant into this market is the PIMA device for PoCT of T-helper cells known as CD4 counts. Measurement of these are an essential feature of guiding antiretroviral therapy for HIV and monitoring the course of immunosuppression. From a design perspective the PIMA is another example of a device that employs the same technology used in laboratory instruments, namely imaging and cell counting, but repackaged into a smaller, more compact instrument which is simpler to operate and can be used to measure CD4 counts on a small sample of whole blood at the point of care. All the required reagents, including labelled antibodies to CD3 and CD4 antigens, are contained in a small disposable cartridge to which is added 5 μL of whole blood, with the cartridge then being inserted into a small battery-powered desktop instrument. After insertion the sample is pumped into the incubation compartment where it interacts with the reagents for a defined period of time after which the stained sample moves to the reading compartment where the fluorescent signal is detected by a CCD camera. Following computational analysis, the results are displayed on the screen. Several evaluations, both in the developed and developing world, show that this instrument, in the hands of non-laboratory trained personnel, provides results of quality comparable to those obtained from the central laboratory.^42,43 By providing CD4 results at the point of care there is the potential to significantly improve treatment by offering prompt immunotherapy to those in need.

Emerging PoCT Technologies

Many of the technologies currently being used for PoCT were generally devised decades ago and have not changed in a fundamental sense in the intervening period. That is not to say that these technologies have not been improved incrementally through advances in materials, electronics and computing technologies amongst other developments. But new technologies that offer substantially different capabilities and potential, and have made the difficult transition to market, are few and far between. Below we briefly review those that may be commercial propositions in the coming decade and two that are already commercially available.

Two decades ago there was much discussion about a concept called Lab on a Chip (LOC) and the view was that this would become the dominant PoCT technology in the future. The development of LOC, also sometimes referred to as microchips, grew from the microelectronics industry through techniques of miniaturisation and microfabrication.⁹ Such devices have been defined as ones that perform analysis at microscopic scales i.e. 1–500 µm and incorporate microfilters, microchannels, microarrays, micropumps, microvalves and bioelectronics chips.⁹ The microchip integrates into one reaction cell all the processes associated with analysis from the placement of the sample into the chip to the analysis itself. Microchips can be fabricated from silicon, glass or polymer. Details of this technology are not often found in the mainstream clinical laboratory literature but Bazydlo et al. provide an interesting and detailed account of this important area.¹⁴ However despite the continuing advances taking place in silicon chip manufacturing contributing to the exponential increase in computing power from such chips, so-called Moore’s Law,⁴⁴ these advances have not yet translated into many commercial devices for PoCT. The notable exception is the i-STAT technology, but there remains considerable interest and research in this area. There is even a dedicated journal called Lab on a Chip, and as will be discussed below, potential products have been described in the research literature. Translating experimental concepts into commercial devices has proved difficult for many reasons, not the least of which is cost.

Due to slow commercialisation of the LOC concept, lateral flow strip (LFS) technology has continued to dominate the PoCT market, as seen in many of the products described previously. But lateral flow strips have a number of limitations and these are shown in Table 5 and described in relation to particular components of the strip by Wong et al.⁴⁵ Collectively these limitations result in two major disadvantages which make it difficult for LFS to meet the needs of some PoCT applications. One such need is so-called multiplexing, namely the ability to measure multiple analytes on the same strip. While this is not impossible using LFS it remains difficult to do for more than two or three analytes.

Table 5.

Typical components of a lateral flow strip, the materials from which they are made and the potential problems that need to be overcome in their design (modified from ref.60).

Component	Function	Material	Potential problems
Sample separation pad	Convert the analytical sample to one suitable for analysis	Cotton linter, glass fibre, rayon	Flooding with excess sample, cannot always filter out contaminants
Conjugate pad	Couples the sample analyte to the measurement conjugate	Glass fibre, polyesters	Variation in uptake, inconsistent binding and release of conjugate
Reaction membrane	Acts as capturing mechanism and forms a visible line when analyte present or absent	Nitrocellulose, nylon	Inconsistent flow characteristics, protein incompatibility to bind with nitrocellulose
Waste reservoir	Acts as sink for excessive sample	Cotton linter	Overflow into assay area
Backing	Provide rigidity and enable easy handling	Polystyrene	Can cause variations in run time and appearance of fluid front
Device housing or Cassette	Protect the properties and features of the device	Plastic	Overflow of sample into cassette

The second major problem for LFS technology is limited sensitivity and this has been brought into focus by the need for better technology for infectious disease testing, particularly in the developing world.⁶ While there have been incremental improvements in the performance of strip tests for various infectious diseases, a recent review shows that many do not meet the required sensitivity to be practically useful.⁴⁶ For example, with tuberculosis testing, several studies have shown that current strip technology is neither accurate nor cost effective and the WHO has recommended they should not be used in individuals suspected of active pulmonary or extra-pulmonary tuberculosis.⁴⁷ With increasing focus on the link between infectious disease and poverty in the developing world, there is a commensurate effort to develop better alternatives to strip tests for infectious disease testing and improve global health.⁵ It is also important to emphasise that there is a considerable burden of infectious disease in the developed world, including respiratory and sexually transmitted infections, which also require better PoCT technology for faster diagnosis and treatment.

The search for better technologies has been directed down two interrelated pathways. One is the continuing efforts to develop the LOC concept, particularly for detection of nucleic acids, and this reflects the general trend in microbiology and infectious disease of serological assays being replaced by molecular testing. The second is the development of paper-based analytical devices which have the advantage that paper can be machined in similar ways to silicon but it is much cheaper, an important consideration for devices in the developing world.

Although progress with developing the LOC concept has been slow, commercial devices using molecular assays to detect infectious agents are starting to appear. There are many challenges associated with developing point of care molecular tests, not the least of which is integrating sample preparation with the other analytical sub-processes to produce a complete analytical process requiring no operator involvement i.e. ‘sample in – result out’.⁴⁸ Up until recently, all devices required laboratory-like facilities to prepare the sample and avoid contamination, which is a major barrier to using the device at the point of care. One device, the Cepheid GeneXpert system described in more detail below, has managed to overcome this integration challenge although it might be argued that it is not really a microfluidic device.

Another challenge is how to provide real-time amplification of the sample. PCR requires a source of heat and therefore power, which may not be available in many developing world point of care locations. Thus various isothermal and enzyme-free amplification techniques have been developed such as loop-mediated isothermal amplification (LAMP) and rolling circle amplification (RCA).⁴⁹

The third major component of point-of-care molecular devices is the type of detection and read-out modality. Optical detection methods include absorbance, light-scattering and fluorescence based measurements while some designs rely on electrochemical measurements.⁴⁹ Of perhaps greater significance is how the detected signal is presented to the user, particularly one using the device in a remote location in the developing world. Several applications have been described of using mobile phones and other telemedicine applications in order to link the person conducting the test to remote expertise which can interpret the result and guide treatment.⁵⁰

The GeneXpert system might be seen more as a system to automate PCR-based assays rather than a PoCT device. But the ability of this relatively small bench-top device to perform real-time quantitative PCR in approximately 90 min with minimal operator interaction means that it offers the potential to perform rapid molecular testing in situations where the need for results is urgent.⁵¹ The system uses single-use cartridges that each contain multiple chambers that hold the sample, various purification and elution buffers, and all the PCR reagents including enzymes; in addition all waste is retained within the cartridge. Attached to the cartridge is a PCR tube around which are heating and cooling tubes together with optical blocks that perform the amplification and fluorescence based detection of the products. Within the cartridge is a syringe body designed in such a way to virtually eliminate sample contamination. Through a series of valves in the syringe, sample is moved through the various stages of the PCR process using the reagents stored in the cartridge, and culminating in real-time detection of the amplified products.⁵²

Several evaluations have been performed of the GeneXpert system for a number of clinical applications including infectious and sexually transmitted diseases, using various sample types. Thus Spencer et al. showed the GeneXpert system had 100% sensitivity and specificity for methicillin-resistant Staphylococcous aureous in paediatric specimens,⁵³ while Buchan et al. tested stool specimens and demonstrated near perfect sensitivity and specificity for Clostridium difficile detection.⁵⁴ An economic-based analysis of tuberculosis testing showed that the GeneXpert system was also cost-effective.⁵⁵

Another commercially available PoCT device for DNA testing is the Spartan RX™ platform which detects mutations in the CYP2C19 gene. These mutations cause reduced response to clopidogrel, an anti-platelet aggregation drug given to patients undergoing percutaneous coronary interventions (PCI), and this can result in adverse consequences. PCIs are often performed urgently, hence the potential benefits for rapid PoCT to detect mutations in the CYP2C19 gene and alternative drug therapy with better outcomes. The Spartan desktop instrument can extract and analyse DNA from a buccal swab. The latter is placed into a cartridge which is then inserted into the instrument and results are available within one hour.⁵⁶ There is limited data on the outcomes of using this test but a recent pilot study of 137 consecutive subjects with acute coronary syndrome and undergoing PCI, on whom the Spartan test was performed, suggests that the test was useful in identifying patients at risk of a poor response to clopidogrel.⁵⁷

Another promising technology for molecular and other types of testing is that being developed by Atlas Genetics. The system comprises a test-specific disposable cartridge containing all the necessary reagents to perform the test. The cartridge is designed to be run on the Atlas io™ reader. Following inoculation of the specimen into the cartridge and insertion of the cartridge into the reader, the test is performed under complete instrument control with no further user input required. The test is based on three fundamental processes that occur sequentially on the test cartridge: (i) DNA extraction; (ii) amplification using PCR; and (iii) detection of specific DNA using an electrochemically labelled probe. The Atlas io™ reader uses pneumatic control to transport the specimen around the various functions within the test cartridge. All reagents are present on the test cartridge as either liquid reagents which are released under reader control, or as dried reagents deposited into the appropriate chambers within the cartridge. These are reconstituted as the specimen enters the chamber containing the deposited reagent. The system is capable of multiplexing by virtue of a number of independent fluidic channels and the opportunity to use a number of electrochemical labels.^58,59

Much has been published in the research literature on the development of paper-based PoCT devices including a comprehensive review by Yetisen et al.⁶⁰ At this point it is important to distinguish nitrocellulose which is in lateral flow strips from paper-based devices. Nitrocellulose is a hydrophobic material which is good for binding to proteins but the hydrophobicity has other disadvantages which paper can potentially overcome including being able to be patterned to form microfluidic devices that are capable of multiplexing. Yetisen et al. identifies four capabilities of such paper devices:

1. Sample can be dispensed into multiple spatially-segregated areas which enable simultaneous assays to be performed on the same device.

2. Samples can move by capillary action so no pumps are needed.

3. Only small sample volumes are required.

4. No hazardous waste as the device can be incinerated.

Many groups including organisations such as ‘Diagnostics for All’ are developing such devices for the developing world primarily because paper is a significantly cheaper material than materials used in lateral flow strips. In addition, the potential applications are wider than the infectious disease area with one group developing a paper microfluidics device the size of a postage stamp for measurement of three liver functions tests – so-called ‘Lab on a Stamp’.⁶¹ While the potential of this technology is high, Yetison et al. comment that no mature platform for this technology has yet appeared, let alone commercial applications.⁶⁰ However Pollock et al. describe encouraging results from a field evaluation of a paper-based device to measure alanine transaminase. ⁶²

Conclusions

The volume of testing performed outside of the conventional laboratory will undoubtedly grow, driven by the need to deliver care closer to the patient. In the last two decades the menu of tests that can be performed using PoCT devices has expanded considerably. This has been achieved largely using well-established technologies such as lateral flow strips but incremental improvements have taken place in many areas so that devices are easier to use, they are less prone to errors, they are more compact or smaller, and in many cases the analytical performance is now more than sufficient for clinical purposes. It is likely that such incremental improvements with existing technologies will continue as the spin off from the continual miniaturisation of electronics and increased computing power that are taking place in other markets such as consumer goods. Examples of technologies that look promising for the future include contact lens glucose sensors,⁶³ tattoo-based sensors⁶⁴ and smart holograms,⁶⁵ all of which are very patient-centred in their configuration.

Yet there are clearly areas where new technologies are needed in order to deliver the required analytical performance. Infectious disease is one such area where existing LFS technology is not sufficiently sensitive and molecular techniques are seen as the future, both in the developed and developing world. PoCT devices that use PCR to detect a number of different pathogens are just starting to appear and in the next few years we are likely to see a number of these reach commercialisation and be used at least within western healthcare markets. Given that the analytical performance of these devices is well proven, the remaining challenge will be to see if they can be produced at a price which will enable them to have an impact on dealing with infectious disease in developing countries.

The final important issue is the ever important need to look beyond the technology and also consider how the result is actually going to change the outcome for the patient – test results alone are useless. This review has concentrated on the analytical part of the technology and has not considered how the result will be delivered to those who will use it to make decisions about treatment etc. This post-analytical part of the testing process is dealt with in the accompanying review of Informatics by Jones et al.⁶⁶ But it is important to emphasise here that post-analytical processes are as critical to the whole value proposition of testing as the measurement technology itself. It is also probably fair to say that its importance has until recently been underestimated and this has contributed to the criticism that much new technology does not deliver benefits in proportion to the investment. This is particularly important for PoCT devices because there has been a preoccupation with the expense of a point of care test compared to one performed in the central laboratory. Yet, as several studies have shown, when economic studies of PoCT are performed that consider the complete testing process, including the patient outcomes, the PoCT value proposition is shown to be favourable compared to the central testing model.^55,67,68 Successful innovation and the adoption of devices which improve patient outcomes in a cost effective way has to include the process and economic changes that are part of the clinical and cost effectiveness evidence – particularly the disinvestment of redundant resources.