FIELD OF THE INVENTION

[0001] This disclosure relates to microfluidic devices suitable for concentrating pathogens in fluid samples with low bacteria concentration.

BACKGROUND

[0002] Pathogen identification is a necessity for accurate prescription of treatments. The capability of rapid diagnosis of pathogens within hours is critical to saving patients in life-threatening conditions such as sepsis. However, early diagnosis of pathogen infection faces the challenges associated with a low number of pathogens in clinical samples. For life-threatening conditions, such as sepsis and pneumonia, patients start to show symptoms at a low bacteria concentration of 1 to 10 colony forming units (CFU)/mL in either bloodstream or pulmonary effusion. Such a low concentration of cells is far below the requirement for diagnosis gold standards, e.g. matrix-assisted laser desorption/ ionization time-of-flight (MALDI-TOF) mass spectrometry requires > 10 ⁶ CFU/mL. Thus, sample culture (> 24 hours) to populate potential bacteria into colonies is necessary for most current practices. However, false culturenegative (i.e. no bacteria colonies formed) are a common problem since the same culture condition can’t satisfy different growth requirements by different bacteria (e.g. aerobic and anaerobic bacteria). Genetic analysis, such as gene arrays and whole-genome sequencing, are culture-independent approaches. Although genetic analysis methods can identify pathogens within hours when dealing with a low number of cells, the amplification procedures necessary to generate enough genetic materials are costly.

[0003] Advances in single-cell technologies has opened a new avenue to study cells with unprecedented speed, throughput, and information. In recent years, Raman spectroscopy has emerged as a non-invasive and label-free technology to study microorganisms. A single cell Raman spectrum (SCRS) contains >1500 Raman bands, providing intrinsic profiles of nucleic acids, protein, carbohydrates and lipids of the cell, which enables characterization of different cell types and their physiological states. Raman spectroscopy coupled

SUBSTITUTE SHEET (RULE 26) with deuterium isotope probing has become a generic method to evaluate cell metabolic activity and thus applied in antibiotic susceptibility tests. Although single-cell approaches remove the need for lengthy culture, enriching low levels of pathogens remains a prerequisite.

[0004] Bacterial cells are ~1 m in size and difficult to separate from other components in clinical samples with conventional bulk centrifugation. Directly isolating low levels of bacteria cells in clinical samples have been reported using ligands modified beads/substrates to bind bacteria. These are effective if the pathogen type is known; however, in most clinical diagnoses, the profile of pathogens is unknown.

[0005] Microfluidic technologies, including dielectrophoresis (DEP) technologies, have recently demonstrated their ability to sort and enrich bacteria cells directly from body fluids. This ligand-free isolation has generic applicability and is well-suited for enriching pathogens without prior knowledge of pathogen type.

[0006] For cell enrichment, positive-DEP (pDEP) design is usually used to avoid heating issues and provide high DEP force. However, pDEP requires low media conductivity (<10 mS/m), while the conductivity of body fluids ranges from 0.1 -3.4 S/m with an average conductivity of 2.15 S/m. To date, most of the work on cell manipulation using pDEP requires off-chip pre-treatment (such as centrifugation and washing steps) to reduce the conductivity of samples.

[0007] Slouka, Z. et al, “Microfluidic Systems with Ion-Selective Membranes”, in Annual Review of Analytical Chemistry, Vol 7, R.G. Cooks and J.E.

Pemberton, Editors. 2014. p. 317-335 proposed fabricating on-chip ionexchange membranes within a microfluidic chip. The chip was then used to reduce the conductivity of samples, however the system can only process a low sample volume. In addition, fabricating on-chip membranes is technically challenging and variable.

[0008] D'Amico, L., et al., “Isolation and concentration of bacteria from blood using microfluidic membraneless dialysis and dielectrophoresis”, Lab on a chip, 2017, p. 1340-1348 reported using a membraneless H-filter to dilute the salts in a blood sample followed by bacteria capture using the pDEP force. However, this approach has low desalting efficiency, relatively low bacteria isolating rate (<80%) and low process throughput (0.6 mL/hour). The isolation rate was only improved to 90% by using a 2x10 ⁷ CFU/mL initial concentration. The high bacteria concentration required for adequate isolations renders this approach unsuitable for samples with low bacteria concentration, for example in patients with early stage symptoms of life-threatening conditions as discussed above.

SUMMARY

[0009] The foregoing summary is only intended to provide a brief introduction to selected features that are described in greater detail below in the detailed description. As such, this summary is not intended to identify, represent, or highlight features believed to be key or essential to the claimed subject matter. Furthermore, this summary is not intended to be used as an aid in determining the scope of the claimed subject matter.

[0010] One aspect of this disclosure relates to a microfluidic device comprising a membrane, a first channel and a second channel. The first and second channels are provided on opposing sides of the membrane and in registration with each other. A first pump is configured to provide a flow of a fluid sample along the first channel in a first direction. A second pump is configured to provide a flow of a diluent along the second channel in a second direction opposite to the first direction such that the diluent flows in the reverse direction to the fluid sample. A third channel is provided, having a first end and a second end remote from the first end, the first end in communication with the first channel to receive the fluid sample therefrom. A plurality of electrodes is provided in the third channel and configured to form a dielectrophoresis capture region in the third channel. An opening is provided at the second end of the third channel, and a metallic surface provided at the second end of the third channel opposite to the opening.

[0011] The details of one or more embodiments of the subject matter described in this specification are set forth in the accompanying drawings and the description below. Other features, aspects, and advantages of the subject matter will become apparent from the description, the drawings, and the claims. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Various exemplary embodiments are illustrated by way of example, and not by way of limitation, in the figures of the accompanying drawings in which like reference numerals refer to similar elements and in which:

[0013] FIG. 1a and FIG. 1 b are simplified functional diagrams of a microfluidic device according to this disclosure;

[0014] FIG. 2 is a top view of a microfluidic device according to this disclosure;

[0015] FIG. 3 is a top view of a baseplate of the microfluidic device of FIG. 2;

[0016] FIG. 4 is a view of electrodes forming part of a capture region in the microfluidic device of FIG. 2;

[0017] FIG. 5a and FIG. 5b are cross-section views of part of the baseplate and capture module of the microfluidic device of FIG. 2;

[0018] FIG. 6 is a graph showing the capture efficiency of the microfluidic device of FIG. 2 at various flow rates;

[0019] FIG. 7 is a functional diagram of the microfluidic device of FIG. 2;

[0020] FIG. 8a is an image of electrodes of the microfluidic device of FIG. 2 before starting desalination and cell capture;

[0021] FIG. 8b is an image of the electrodes of FIG. 8a after 10 minutes of desalination and cell capture;

[0022] FIG. 9a is a microscope image of cells captured in the microfluidic device of FIG. 2;

[0023] FIG. 9b is the Raman spectrum of a cell shown in FIG. 9a.

DETAILED DESCRIPTION

[0024] This disclosure relates to microfluidic devices suitable for concentrating pathogens in fluid samples with low concentration. To assist the reader in better understanding the disclosure we first describe a simplified functional diagram as shown in FIG. 1a and FIG. 1 b, prior to describing specific detailed embodiments and examples.

[0025] FIG. 1a shows, in simplified form, a microfluidic device 10 comprising a first channel 12 and a second channel 14. The first and second channels 12, 14 are microfluidic channels. The first and second channels 12, 14 are provided on opposing sides of a membrane 16 and are in registration with each other.

Put another way, in the orientation shown in FIG. 1 , the first channel 12 is directly above the second channel 14 with the membrane 16 separating the first and second channels 12, 14. The membrane 16 may be a membrane microfilter with a suitable pore size or a semi-permeable membrane according to requirements. In the described embodiment, the membrane 16 is a membrane microfilter.

[0026] A first pump (not shown) is configured to provide a flow of a fluid sample along the first channel 12 in a first direction indicated by the arrows.

[0027] A second pump (also not shown) is configured to provide a flow of a diluent along the second channel 14 in a second direction indicated by the arrows. The second direction is opposite to the first direction such that the diluent flows in a reverse direction to the fluid sample.

[0028] The membrane microfilter 16 is configured to enable ions to move from the fluid sample in the first channel 12 into the diluent in the second channel 14 by diffusion through the membrane microfilter 16. Thus, the fluid sample is desalinated as it moves along the first channel 12.

[0029] A third channel 18 is in communication with the first channel 12 to receive the desalinated fluid sample therefrom. A dielectrophoresis (DEP) capture region 20 is provided in the third channel 18. The DEP capture region 20 comprises a plurality of electrodes which, when activated by a control signal, cause cells in the desalinated fluid sample to be attracted to the electrodes and held in place. As the desalinated fluid sample continues to flow past, the number of cells held in place increases, leading to concentration of the cells at the DEP capture region 20. In some embodiments, the DEP capture region 20 and the control signal may be configured to cause cells with specific electrical and/or geometrical properties to be attracted to the electrodes and held in place, while other cells or particles with different electrical or geometrical properties are either pushed away from the electrodes with an opposing force or do not experience sufficient DEP force to be retained against the flow of the fluid sample. Optionally, the geometry of electrodes can be designed to perform electrophoresis.

[0030] FIG. 1 b shows the DEP capture region 20 in simplified form, in which electrodes 270, 280 are shown. The third channel 18 in the vicinity of the DEP capture region 20 is dimensioned according to the flow rate, DEP capture requirements and manufacturing materials/methods. In some embodiments, the third channel 18 has a height, shown as ‘h’ in FIG. 1 b, in the range 20-75 microns and a width (into the page when looking at FIG. 1 b) in the range of 1-3 mm. In some embodiments, the electrodes 270, 280 have a track width and gap between them in the range 20-50 microns. It will be appreciated that DEP field strength reduces as distance from the electrodes increases. The third channel 18 having a height that is comparable to the electrode track width and gap between electrodes results in a DEP field having useful strength throughout the third channel 18, though other configurations are possible. As illustrated in FIG. 1 b it is preferred that the electrodes 270, 280 are formed embedded or recessed into the surrounding substrate so the third channel 18 is free from ‘steps’ or protuberances to which cells may attach and be difficult to remove from.

[0031] Referring again to FIG. 1 a, an opening 22 is provided at an end of the third channel 18 remote from the first channel 12. A metallic surface 24 is provided at the end of the third channel 18 opposite the opening 22. Cells held in place at the DEP capture region 20 can be released by removing the control signal to the plurality of electrodes. The cells will then move with the fluid flow along the third channel 18 towards the opening 22. The concentrated cells can be moved along from the DEP capture region 20 and retained in the vicinity of the opening 22 and metallic surface 24 by turning off the first pump. The fluid together with cells can be retrieved off-chip. Alternatively, the fluid can be allowed to evaporate to leave behind the cells which will then be on the metallic surface 24. The opening 22 can then act as a window to permit inspection and/or identification of the cells by any suitable method, e.g. microscope or other optical imaging, Raman spectroscopy, etc. The metallic surface 24 has a low surface roughness and acts as a mirror for optical imaging and is preferably formed of Nickel which has been found to possess low Raman emission properties, rendering it an excellent background for Raman spectroscopy of the cells. In some embodiments, the metallic surface 24 may be formed of Tin. Optionally, the metallic surface can also be roughened or patterned with plasmonic structures to enhance Raman signal locally, for instance to facilitate surface-enhanced Raman scattering (SERS).

[0032] The first channel 12, the second channel 14 and the membrane microfilter 16 form a membrane-based desalinator. The use of a membrane enables the flow rate of the fluid sample to be controlled independently of the diluent, unlike a membrane-less desalinator.

[0033] It has been found that the rate of fluid flow past the DEP capture region can adversely affect the proportion of cells captured if the flow rate is too high. The membrane-based desalinator allows the flow rate of the fluid sample in the first channel 12 to be at a rate suitable for cell capture in the DEP capture region 20, which in turn enables the third channel 18 to receive the desalinated fluid sample from the first channel 12 such that desalination of the fluid sample and cell capture and concentration is performed at the same time in the microfluidic device. In addition, the use of a membrane 16 ensures that no cells in the fluid sample are ‘lost’ due to flowing out with the diluent, since cells cannot pass through the membrane 16.

[0034] DEP capture performance is very sensitive to the presence of ions in the fluid sample, thus adequate desalination is necessary to ensure cells are captured in the DEP capture region 20. It has been found that to achieve the necessary desalination levels with the fluid sample flow rate at a level suitable for DEP capture, the diluent in the second channel 14 must flow in the opposite direction to the direction the fluid sample flows through the first channel 12.

[0035] The high DEP capture performance combined with desalination efficiency makes the microfluidic device 10 suitable for concentrating pathogens in cases where a fluid sample contains very low bacteria concentration, for instance less than 10 CFU/mL. In addition, the microfluidic device 10 provides a convenient and fast solution, since desalination and cell concentration occur in the same process, reducing the time required to obtain a cell concentration sufficient for pathogen identification. It will be appreciated the device may also be used with fluid samples with pathogen concentrations higher than 10 CFU/mL.

[0036] Having described a simplified functional diagram of the microfluidic device illustrated in FIG. 1 , we now describe more detailed embodiments and examples below.

[0037] Turning now to FIG. 2, a microfluidic device 100 is shown, with like reference numerals denoting like parts to those of the microfluidic device 10.

[0038] The microfluidic device 100 is implemented in a modular configuration with a desalinator module 110 and a capture module 120 mounted to a baseplate 130. The modular configuration may permit, for example, the baseplate 130 and desalinator module 110 to be reused and optionally the capture module may be removed after cell concentration and transferred to a microscope or spectrometer stage.

[0039] The desalinator module 110 comprises the first channel 12, the second channel 14 and the membrane microfilter 16. The first channel 12 and the second channel 14 have a circuitous configuration as seen in FIG. 2 to increase the surface area in contact with the membrane microfilter 16. FIG. 2 is a top view of the microfluidic device 100 in which the first channel 12 is visible, while the second channel 14 is below the first channel 12 and in registration with the first channel 12 and so is not visible.

[0040] The first channel 12 and the second channel 14 are each formed in separate layers of suitable material, such as Poly(methyl methacrylate) “PMMA”, although other plastic materials may be used.

[0041] The membrane microfilter 16 is circular in the configuration shown in FIG. 2 though other shapes may be adopted. The membrane microfilter 16 is sandwiched between the layers in which the first channel 12 and the second channel 14 are formed, as generally shown in FIG. 1. The membrane microfilter 16 may be formed of cellulose acetate with 0.45 micron pores in one implementation, although other suitable materials may be used. A pore size in the range of 0.2 to 0.5 micron is preferred. While smaller pore size is generally preferred, smaller pore size also requires longer channels 12 and 14 to achieve the same desalination performance.

[0042] The first channel 12 has an inlet 170 to receive a sample fluid via a first pump (not shown) and an outlet 190 that connects the first channel 12 to the third channel 18 of the capture module 120. The first pump is configured to provide a flow of a fluid sample along the first channel 12 in a first direction from the inlet 170 to the outlet 190. In the configuration shown in FIG. 2, the outlet 190 connects the first channel 12 to the third channel 18 via an intermediate channel 200. A bleed channel 210 is formed in the baseplate 130 which connects the intermediate channel 200 to a connector 145c. The bleed channel 210 permits removal of air bubbles trapped in the first channel 12 or the intermediate channel 200. In some configurations a sensor, such as a conductivity sensor, may be used to detect the presence of air bubbles and activate a pump connected to connector 145c to remove the air via the bleed channel 210, for instance during priming of the device 100.

[0043] The desalinator module 110 is held in place on the baseplate 130 via a clip 160. When the desalinator module 110 is mounted to the baseplate 130, the second channel 14 is connected at each end to channels 140a, 140b formed in the baseplate 130. The channels 140a, 140b terminate in connectors 145a, 145b on the baseplate 130. A second pump (also not shown) is configured to provide a flow of a diluent along the second channel 14 in an opposite direction to the first direction such that the diluent flows in a reverse direction to the fluid sample. In the configuration shown in FIG. 2, connector 145a is connected to the second pump that provides a flow of diluent to the device 100 and connector 145b is a waste diluent outlet for connection to a suitable waste fluid receptacle or disposal system. The diluent may be any suitable diluent, such as purified water and preferably deionized water although distilled water may also be used. The desalting efficiency of the desalinator module 110 was evaluated using an LB broth with a conductivity of 2.25 S/m - corresponding to a typical clinical urine sample. After passing through the desalinator module 110, the conductivity of the LB broth was reduced by a factor of 2000, from 2.25 S/m to 1 .3 ± 0.12 mS/m, suggesting a high salt removing efficiency. A urine sample spiked with 10 ⁴ CFU/mL E. coli was then tested in the system with the flow rate at 40 mL/min. After passing through the desalinator module 110 the conductivity of the urine sample was reduced to 0.96 ± 0.57 mS/m.

[0044] Gaskets 150 form a seal between the desalinator module 110, the capture module 120 and the baseplate 130 wherever channels interconnect between these elements.

[0045] As discussed above, the presence of the membrane microfilter 16 enables independent control of flow rates through the first channel 12 and the second channel 14. It has been found that DEP capture efficiency of greater than 95% can be achieved by optimizing flow rate and electric field. Flow rates of 10-500 pL/min, can be used. For higher flow rates higher electric field strength is needed to maintain high capture efficiency, typically achieved by higher control signal voltages. For example, at flow rates above 40 pL/min, in the first channel 12 and third channel 18 efficiency reduces below 95 % with a control signal of 20 Volt peak-to-peak 1 MHz, as shown in FIG. 6. Whereas with a control signal of 40 Volts peak-to-peak 1 MHz, flow rates of 200 pL/min can be used and high capture efficiency achieved. In contrast, the flow rate of diluent in the second channel 14 can be 50-2500 pL/min, with 200-500 pL/min being preferred. To balance the pressure across the membrane microfilter 16 the second channel 14 has a larger cross-section than the first channel 12, such as a deeper channel as illustrated in FIG. 1 . It will be understood that while a control signal frequency of 1 MHz has been used in the examples above, other control signal frequencies may be used and this disclosure is not limited to the particular frequency used in the examples above.

[0046] The capture module 120 is held in place on the baseplate 130 via a clip 220. The capture module 120 includes the third channel 18. The capture module 120 may be conveniently sized to correspond with a microscope slide, though other shapes and sizes may be used.

[0047] The third channel 18 has a first end 230 and a second end 240 remote from the first end 230. The first end 230 is in communication with the first channel 12 to receive the fluid sample therefrom. In the configuration shown in FIG. 2 the first end 230 is in communication with the first channel 12 via the intermediate channel 200 to receive the fluid sample therefrom.

[0048] A plurality of electrodes 250 are provided in the third channel 18 and configured to form a DEP capture region 20 in the third channel 18.

[0049] Referring now to FIG. 4, which is a view of region A indicated in FIG. 2, the plurality of electrodes 250 is configured as interdigitated positive electrodes 260 and negative electrodes 270. The positive electrodes 260 are shown in FIG. 4 as dashed lines solely to assist the reader in understanding the drawing - it will be appreciated that the positive electrodes 260 are a continuous circuit to provide an electric field.

[0050] Further, the plurality of electrodes 250 are configured as a first region 20a and a second region 20b. In the first region 20a the plurality of electrodes 250 is configured as a ‘V’ pointing in a direction of flow of fluid in the third channel 18. In the second region 20b the plurality of electrodes 250 is configured as parallel electrodes extending transversely across the third channel 18.

[0051] Returning now to FIG. 2, the positive electrodes 260 and the negative electrodes 270 in the regions 20a and 20b are connected to terminals 280 to receive control signals via the terminals 280. In some implementations, a different control signal is provided to the electrodes 250 in the region 20a to the control signal provided to the electrodes 250 in the region 20b. In other implementations, the same control signal is provided to the electrodes 250 in the regions 20a and 20b. The control signal is AC and may have a time-varying frequency component or a fixed frequency component. Control signals are received from a controller such as a signal generator (not shown).

[0052] The first region 20a is closer to the first end 230 than the second region 20b. When a control signal is applied to the electrodes 260, 270 in the first region 20a, the ‘V’ configuration of the electrodes as shown in FIG. 4 acts to direct cells present in the desalinated fluid sample towards the middle of the third channel 18 via DEP. Directing the cells present in the desalinated fluid sample towards the middle of the third channel 18 assists optical inspection and/or optical detection, such as using an imaging device and/or suitable image analysis software, to see if cells are present and the approximate amount. Such optical inspection/detection is useful for establishing the concentration conditions. For example, in trials using urine we have found the conditions are such that there is high confidence there will be sufficient cells captured and therefore there is no longer a need to inspect region 20a as part of the test procedure. As such, depending on the concentration conditions, regular or routine inspection of the first region 20a may be omitted once confidence is established via inspection.

[0053] When a control signal is applied to the electrodes 260, 270 in the second region 20b, the transverse configuration of the electrodes as shown in FIG. 4 acts to capture cells present in the desalinated fluid sample via DEP. As illustrated in FIG. 6, the DEP capture region 20 has demonstrated capture efficiency, with a 20 Volt peak-to-peak 1 MHz control signal, of over 90% at flow rates at or below 40 pL/min and approaching 100% at flow rates at or below 20 pL/min, thus making the device suitable for fluid samples with low cell concentrations. Using a control signal voltage of 40-50 Volts can achieve over 95% efficiency at a 200 pL/min flow rate. While still higher voltages might be used, it will be understood that excessive voltage can lead to heating effects at the electrodes which may be detrimental, e.g. to the device materials and/or to the health of the cells. Cells in the desalinated fluid sample continue to be attracted to the electrodes 260, 270 and held in place as the desalinated fluid sample flows past, thus concentrating the cells at the second region 20b of the DEP capture region 20. In a similar manner as described above for the first region 20a, optical inspection and/or optical detection may be performed in the second region 20b, in this case to determine whether a sufficient concentration of cells have been captured in the second region 20b. Detection and inspection of the first and second regions 20a, 20b may be performed using the same imaging device. Further, other detection methods besides optical may be used as discussed below.

[0054] In some configurations, a second outlet path 290 is provided in the capture body 120 which is in communication with the third channel 18 after the DEP capture region 20 and proximate to the second end 240 of the third channel 18. The second outlet path 290 is in communication with a sample waste channel 295 formed in the baseplate 130. The sample waste channel 295 terminates at a connector 145d (not labelled in fig 3). A third pump (not shown) is connected to the connector 145d and configured to remove fluid from the third channel 18 via the sample waste channel 295 and linked to the second outlet path 290. That is, the third pump is configured in a pull or suction mode rather than a push or pump mode. It has been found that the use of the third pump in addition to the first pump is beneficial in maintaining a desired flow rate through the third channel 18. In some implementations, the second outlet path 290 may be conveniently used to collect larger quantities of cells, e.g. for antibiotic susceptibility testing.

[0055] Concentration of cells at the DEP capture region 20 may be determined to be complete according to any suitable criteria. In one example embodiment, concentration may be determined using impedance measurements between the positive electrodes 260 and the negative electrodes 270. In another example embodiment, an imaging device may be used to count the cells in the second region 20b, wherein concentration may be determined to be complete once a threshold quantity or density of cells is determined. A window 300 may be provided in the baseplate 130 to avoid the baseplate 130 from distorting imaging, reducing resolution, of the second region 20b. FIG. 8a shows an image of electrodes 260, 270 in a portion of the second region 20b from an imaging device at the start of desalination and cell concentration. FIG. 8b shows an image of the same electrodes 260, 270 after 10 minutes. Cells 310 can be seen in FIG. 8b on the electrodes 260, 270. The scale bar in FIGs. 8a and 8b represents 20 pm.

[0056] In another example, concentration may be determined to be complete once a threshold volume of fluid has passed along the third channel 18, which may be determined by the first or third pumps and/or their controllers. In yet another example, concentration may be determined to be complete once a period of time has elapsed after control signals are applied to the electrodes 260, 270. It has been found that these latter two methods of determining concentration are suitable for samples with high bacteria concentration, such as urine. [0057] Referring now to FIG. 5, which shows the capture body 120 mounted to the baseplate 130, the opening 22 is provided at the second end 240 of the third channel 18. The opening 22 extends generally perpendicularly to the third channel 18. The metallic surface 24 is provided at the second end 240 of the third channel 180 opposite to the opening 22.

[0058] A first outlet path 320 is provided in the baseplate 130 shown in FIG. 3. The first outlet path 320 is in communication with the opening 22 when the capture body 120 is positioned on the baseplate 130. The first outlet path 320 terminates at a connector 145e.

[0059] Once concentration is determined to be complete, the third pump is stopped. The control signals to the electrodes 260, 270 in the DEP capture region 20 are switched off, releasing the captured bacteria/cells. Once released, the cells are pushed with the sample fluid to the second end 240 of the third channel, which can be achieved by controlling the volume of fluid pumped using the first pump after the cells are released. After that, the first and second pumps are stopped which stops the sample flow and diluent flows. The once-captured cells are now in the vicinity of the metallic surface 24 and the opening 22. In some implementations, the electrodes 260, 270 in the DEP capture region 20 have a surface roughness preferably less than 100 nm rms and more preferably 10-30 nm rms, which has been found to aid in releasing captured bacteria/cells.

[0060] The capture body 120 can be removed from the baseplate 130 using the clip 220. FIG. 5b shows the capture body 120 removed from the baseplate 130 and inverted so the opening 22 faces upwards. The cells can be easily pipetted off-chip via the opening 22 for other processes. Alternatively, the cells may be dried in-situ for on-chip detection. As the fluid sample evaporates, the cells are left remaining on the metallic surface 24. The desalination process also functions as an effective “washing” step by removing ions and small molecules, thus preventing crystal formation during the drying process. The opening 22 avoids any interference of device materials on the cell inspection, e.g. if using Raman spectroscopy, since inspection can be performed through the opening 22. Both avoidance of crystals and avoidance of device materials are advantageous for acquiring single-cell Raman spectra of high signal-to- noise ratios, allowing accurate classification of pathogens within a few minutes. It is preferred that the metallic surface 24 is formed of Nickel, which has been found to have around 40 times less Raman emissions than a typical Raman- emission from a cell and thus provides an excellent background for Raman spectroscopy. The capture body 120 may conveniently be dimensioned to correspond with a microscope slide to provide easy use in inspection instruments. The capture body 120 may be removed from the baseplate 130, turned over and repositioned on the baseplate 130 with the outlet 22 facing away from the baseplate 130 or placed on the microscope stage of an optical or Raman microscope (not shown).

[0061] FIG. 9a shows an image taken through the opening 22 by a Raman microscope which clearly shows cells dried on the metallic surface 24. The scale bar represents 20 pm.

[0062] FIG. 9b is an example Raman spectra of a cell in FIG. 9a. In some implementations, a processing module (not shown) may analyse the Raman spectra obtained through the opening 22 and output an identification classification of the cells. In one configuration, the processing module may include a database (also not shown) of Raman spectra of known pathogens which is compared to the Raman spectra of the cells to provide the identification classification. In another configuration, the processing module may include a machine learning classifier that has been trained using a set of Raman spectra of known pathogens. The machine learning classifier may take the Raman spectra of the cells as an input and provide as an output an identification classification of the cells.

[0063] Referring now to FIG. 7, an example functional diagram of the microfluidic device of FIG. 2 is shown. For clarity of the illustration the desalinator module 110 is shown separately to the capture body 120, though it should be appreciated that these may be mounted to the baseplate 130 as described above. As shown in FIG. 7, a function generator 400 provides control signals to electrodes in the DEP capture region 20. The function generator 400 may be controlled by a control system (not shown) which controls operation of the function generator 400 based on determination of concentration of cell in the DEP capture region 20 as described above. For this purpose, the control system may include an optical detector such as an imaging device and suitable image analysis software, or alternatively may include impedance measuring systems as mentioned above. The first pump, the second pump and the third pump are shown in FIG. 7 as 410, 420 and 430 respectively. Arrows shown in FIG. 7 indicate the direction each pump 410, 420 and 430 is configured to operate. The pumps 410, 420 and 430 may each be syringe pumps in some embodiments.

[0064] In some implementations, the desalinator module 110 and the capture module 120 may be formed integrally. In some implementations, the desalinator module 110 may be integrally formed with the baseplate 130.

[0065] Particular embodiments of the invention have been described. The separation of various system components in the embodiments described above should not be understood as requiring such separation in all embodiments, and it should be understood that any described program components and systems may generally be integrated together in a single product or packaged into multiple products.