WO2006119806A1

WO2006119806A1 - Fluid cell with sheath flow

Info

Publication number: WO2006119806A1
Application number: PCT/EP2005/053050
Authority: WO
Inventors: Klaus Witt; Tom Van De Goor
Original assignee: Agilent Technologies, Inc.
Priority date: 2005-05-13
Filing date: 2005-06-29
Publication date: 2006-11-16
Also published as: DE112005003572T5

Abstract

A fluid cell is described, with the fluid cell comprising a first inlet adapted for providing a sample fluid to the fluid cell, a second inlet adapted for providing a sheath fluid to the fluid cell, and one or several outlets. A sample flow of sample fluid is established between the first inlet and one or more of the one or several outlets, and a sheath flow of sheath fluid is established between the second inlet and at least one of the one or several outlets. Said sheath flow is adapted for sheathing the sample flow, at least in a part of the fluid cell. The fluid cell further comprises a receiver adapted for receiving a response signal in response to a stimulus signal coupled to the fluid cell.

Description

FLUID CELL WITH SHEATH FLOW

BACKGROUND ART

[0001] The present invention relates to fluid cells e.g. in a fluid separation system.

[0002] When determining properties of a sample fluid in a detection cell, there often arises a need for confining the sample fluid to a predefined portion of the detection cell. For example, it might be advantageous to confine the sample fluid to a region where detection actually takes place. For example, when determining properties of cells or beads contained in sample fluid, measurement accuracy might be increased by mechanically restricting the flow of sample fluid to a predefined portion of the detection cell.

DISCLOSURE

[0003] It is an object of the invention to provide to provide an improved fluid cell. The object is solved by the independent claim(s). Preferred embodiments are shown by the dependent claim(s).

[0004] According to embodiments of the present invention, a fluid cell comprises a first inlet adapted for providing a sample fluid to the fluid cell, a second inlet adapted for providing a sheath fluid to the fluid cell, and one or several outlets. A sample flow of sample fluid is established between the first inlet and one of the one or several outlets, and a sheath flow of sheath fluid is established between the second inlet and at least one of the one or several outlets, with the sheath flow being adapted for sheathing the sample flow, at least in a part of the fluid cell. The fluid cell further comprises a receiver adapted for receiving a response signal in response to a stimulus signal coupled to the fluid cell.

[0005] A sample flow is set up by supplying sample fluid to the first inlet, and by drawing off the sample fluid via one of the one or several outlets. Furthermore, by supplying a sheath fluid to the second inlet and drawing off the sheath fluid via at least one of the one or several outlets, a sheath flow is set up. Flow may e.g. be driven by applying vacuum on the outlet side or by applying pressure at the inlet side. Also a pressure differential, e.g. different heights of liquid at inlet and outlet or evaporation at the outlet, or an electromobilization through electroendosmosis may be used. The sheath flow is adapted for confining the sample flow of sample fluid to a predefined portion of the fluid cell. When coupling a stimulus signal to the fluid cell, and receiving a response signal in response thereto, one or more properties of the sample fluid may be derived from the response signal. For example, the flow of sample fluid might be confined to a predefined region of the fluid cell, with the stimulus signal being applied to this predefined region. By means of the one or more sheath flows, the flow of sample fluid might e.g. be restricted to the area of the detection cell where detection actually takes place.

[0006] Furthermore, by sheathing the flow of sample fluid, it can be achieved that direct contact between the sample fluid and the fluid cell's sidewalls is avoided or at least reduced. As a consequence, macromolecules, cells, beads or any other species contained in the sample fluid may no longer adhere to the fluid cell's sidewalls, and contamination of the sidewalls is reduced. Furthermore, plugging of the fluid cell's detection channel, which has frequently occurred in prior art detection cells, can be avoided as well. By providing for one or more sheath flows, the sample flow is kept away from the fluid cell's sidewalls, and a well-defined flow geometry is established. As a result, the fluid cell's reliability is improved, and with regard to the response signal, measurement accuracy is improved.

[0007] According to a preferred embodiment, both the first inlet and the second inlet are located in the vicinity of the fluid cell's first end. According to another preferred embodiment, the one or several outlets are located in the vicinity of the fluid cell's second end opposite to the first end. Accordingly, the (one or more) sheath flows and the sample flow substantially flow in parallel from the fluid cell's first end to the second end.

[0008] In a preferred embodiment, the stimulus signal is an electrical signal or an electromagnetic signal. By receiving a response signal in response to the electrical or electromagnetic stimulus signal, electrical or electromagnetic properties of the fluid contained in the fluid cell can be derived. Alternatively, the stimulus signal might be an optical signal, in order to determine an optical property of the fluid in the fluid cell.

[0009] In a preferred embodiment, a first electrode is located in the area of the fluid cell's first end or in the area of the fluid cell's second end, in order to couple an electrical or electromagnetic stimulus signal to the fluid cell. Furthermore, in another preferred embodiment, a second electrode is located in the region of the fluid cell's second end or in the area of the fluid cell's first end, with the second electrode being adapted for receiving an electrical or electromagnetic response signal. In this embodiment, the properties of the fluid contained in the fluid cell are determined across the detection channel's length.

[0010] According to another preferred embodiment, the sheath fluid and the sample fluid flow in direct contact to one another. As long as lateral diffusion remains small, the sheath fluid and the sample fluid do not mix significantly.

[0011] In a preferred embodiment, the fluid cell comprises a determination unit adapted for converting the response signal into one or more electrical properties of the sample fluid. For example, the determination unit might compare the magnitude and/or the phase of the response signal with the stimulus signal's magnitude and/or the stimulus signal's phase.

[0012] In a preferred embodiment, both the stimulus signal and the response signal are AC signals. When detecting an AC response signal, both the response signal's magnitude and its phase shift provide information about the sample fluid's electrical properties. Furthermore, additional information might e.g. be obtained by varying the AC signal's frequency. AC signals may be capacitively coupled to the fluid and coupled out of the fluid contained in the fluid cell.

[0013] In a preferred embodiment, the electrical properly derived from the response signal is at least one of: conductivity, complex conductivity, impedance, resistance, reactance, relative permittivity.

[0014] According to another preferred embodiment of the invention, the sample fluid's conductivity exceeds the conductivity of the sheath fluid. In this embodiment, the electrical properties of the fluid contained in the fluid cell mainly depend on the sample fluid's properties. In fact, the flow of conductive sample fluid can be considered as a "liquid wire". In this embodiment, the presence of the sheath flow does not significantly alter the sample flow's electrical behaviour.

[0015] In a preferred embodiment, the length of the fluid cell is short enough to make sure that the sheath fluid and the sample fluid substantially do not mix. The amount of lateral diffusion depends on the time interval required for traversing the fluid cell. This time interval might e.g. depend on the fluids' respective velocities, and on the length of the fluid cell. If the length of the fluid cell is sufficiently short, the amount of lateral diffusion will be rather small. Preferably, the length of the fluid cell is so small that the sample flow and the sheath flow remain substantially phase separated over the length of the fluid cell. Preferably, the length of the fluid cell is between 1 μm and 1800 μm, and further preferably, the length of the fluid cell is between 10 μm and 300 μm.

[0016] According to a preferred embodiment, at least one of the first electrode and the second electrode is in direct contact with the sample fluid. By directly contacting the sample fluid, highly sensitive measurements are possible.

[0017] In an alternative embodiment, an AC signal is capacitively coupled to the tifluid cell via the first electrode. In another preferred embodiment, an AC response signal is capacitively received via the second electrode. In these embodiments, any direct contact between an electrode and the fluid contained in the sample cell, which might give rise to undesired effects, is avoided.

[0018] In a preferred embodiment, both the sample fluid and the sheath fluid are drained off via one common outlet. In an alternative embodiment, the fluid cell comprises a first outlet adapted for draining off primarily the sample fluid and one or more second outlets adapted for draining off primarily the sheath fluid. In this embodiment, the sample fluid obtained at the first outlet might e.g. be subjected to further analysis. Furthermore, by individually controlling the sub-atmospheric pressures applied to the first outlet and to the one or more second outlets, the respective flow rates of the sample flow and of the sheath flow(s) may be controlled separately. [0019] In a preferred embodiment, a single-sided squeezing of the sample flow is accomplished, with the sample flow being confined by means of one single sheath flow located on one side of the sample flow. In an alternative embodiment, a two-sided squeezing of the sample flow is accomplished by means of two sheath flows located on the right-hand side and on the left-hand side of the sample flow. In yet another embodiment, the sheath flow surrounds the sample flow in a way that an all-sided squeezing of the sample flow is accomplished.

[0020] In a preferred embodiment, the fluid cell is used for at least one of counting and analyzing cells, beads, or a mixture of both contained in the sample fluid. Whenever a cell or bead passes the detection channel, the presence of the cell or bead is indicated by a corresponding change of the electric, electromagnetic or optical response signal. For example, when the flow rate of the sample flow is known, the number of cells or beads per unit volume can be determined. By introducing one or more sheath flows, it is made sure that the cells or beads do not adhere to the side walls, and plugging of the detection channel is avoided, even though the sample stream is confined to an area as small as the sample cells or beads.

[0021 ] According to a preferred embodiment, the flow of highly conductive sample fluid acts as a liquid wire adapted for contacting cells or beads as they pass through the detection channel. This allows determining the electrical properties of a cell or bead in the detection channel. The conductivity of the sheath fluid is much smaller than the sample fluid's conductivity, and for this reason, the one or more sheath flows act as insulators and do not significantly affect the measured electrical properties.

[0022] In a preferred embodiment, an electrical property is modulated whenever a cell passes through the detection channel. In a further preferred embodiment, the determination unit is adapted for analyzing the changes of the respective electrical property. In another preferred embodiment, the determination unit performs a spectral analysis of the response signal. Thus, frequency components absorbed by the cells, or e.g. by functionalized beads, can be detected. In general, absorption of a certain frequency component corresponds to an excitation, e.g. to a rotational degree of freedom. Spectral analysis of the response signal gives a detailed picture of the moieties contained in the sample fluid.

[0023] In a preferred embodiment, the fluid cell's inner surface is coated with an anti-adhesive, like e.g. PEG (polyethylene glycol).

[0024] In another preferred embodiment, the fluid cell further comprises a light source and an optical detection unit. The light source provides a stimulus signal to the fluid contained in the fluid ceil, and the optical detection unit is adapted for analyzing an optical response signal obtained in response to the stimulus signal. For example, the intensity of transmitted light, or the intensity of fluorescence light might be detected, or a beam deflection might indicate refractive index changes.

[0025] According to another preferred embodiment, fluorescence intensity is evaluated each time a measured electrical property indicates that a cell or bead passes through the fluid cell. In this embodiment, changes of a detected electrical property trigger the evaluation of fluorescence intensity. For each cell or bead, it is made sure that fluorescence intensity is determined exactly at the point of time when the cell or bead passes the fluorescence detection unit. By combining the evaluation of electrical response signals with fluorescence intensity detection, a precise monitoring of the individual cells¹ or beads' fluorescence intensity is achieved. Doublets, multiplets or pairs can be distinguished and excluded for clear interpretation of authentic populations.

[0026] According to another embodiment, the electrical property is evaluated each time the fluorescence detection unit indicates that a cell or bead passes through the fluid cell.

[0027] According to yet another preferred embodiment of the invention, the fluid cell might be realized as part of a microfluidic chip device. Further preferably, for manufacturing the microfluidic chip device, suitable techniques such as e.g. laser ablation, hot embossing, etching, micromolding are used. In a preferred embodiment, the microfluidic chip device is realized as a multilayer structure comprising two or more layers. Each of the layers may be microstructured separately before assembly.

[0028] A fluid separation system according to embodiments of the present invention may comprise one or more of: a fluid delivery system, a separation device for separating components of the fluid delivered by the fluid delivery system, and a fluid cell as described above for at least one of analyzing and detecting components of the fluid separated by the separation device.

[0029] The separation device might e.g. be a liquid chromatography or an electrophoresis column. By sheathing the flow of sample fluid obtained from the separation device, measurement accuracy of the fluid cell can be improved. By modulating the pressure values applied to the fluid cell's one or more outlet channels, sample components like e.g. cells or beads can be sorted and made available as homogenous populations.

[0030] The invention can be partly or entirely embodied or supported by one or more suitable software programs, which can be stored on or otherwise provided by any kind of data carrier, and which might be executed in or by any suitable data processing unit. Software programs or routines are preferably applied for controlling operation of the fluid cell.

BRIEF DESCRIPTION OF DRAWINGS

[0031 ] Other objects and many of the attendant advantages of embodiments of the present invention will be readily appreciated and become better understood by reference to the following more detailed description of preferred embodiments in connection with the accompanied drawing(s). Features that are substantially or functionally equal or similar will be referred to with the same reference sign(s).

[0032] Fig. 1 shows a fluid cell according to embodiments of the present invention;

[0033] Fig. 2 depicts a fluid cell having a common outlet;

[0034] Fig. 3 shows a fluid cell comprising an optical detection set-up;

[0035] Fig. 4 shows a functionalized polymeric bead;

[0036] Fig. 5 illustrates the parameters that affect flow geometry; [0037] Fig.6 shows an implementation for realizing a two-sided squeezing of a flow of sample fluid; and

[0038] Fig.7 gives an implementation for realizing an all-sided squeezing of a flow of sample fluid.

[0039] Figs. 1 A, 1 B and 1 C show a detection cell according to embodiments of the present invention. Fig. 1 A gives a top view, Fig. 1 B shows a perspective view, and Fig. 1C shows a cross section of the detection cell. Via a first inlet 1, a flow 2 of sample fluid is supplied to the detection channel 3. Simultaneously, via a second inlet 4, a sheath flow 5 of sheath fluid is provided, and via another second inlet 6, another sheath flow 7 of sheath fluid is supplied. The length of the detection channel 3 is chosen such that the sheath flow 5, the flow 2 of sample fluid, and the sheath flow 7 substantially do not mix. Over the length of the detection channel 3, the sheath fluid and the sample fluid remain substantially phase separated.

[0040] The sample fluid might e.g. comprise a certain concentration of cells 8, 9. In an alternative embodiment, the sample fluid might comprise a certain amount of polymeric beads. Preferably, the surface of these beads has been functionalized, in order to react with some class of chemical compounds. The detection cell further comprises first outlet 10 adapted for drawing off the flow 2 of sample fluid, a second outlet 11 adapted for drawing off the sheath flow 5, and another second outlet 12 adapted for drawing off the sheath flow 7.

[0041] Fig. 1 B shows how the sheath flows 5, 7 flowing on the right-hand side and the left-hand side of the sample fluid under appropriate flow or pressure conditions confine the flow 2 of sample fluid to a center portion 13 of the detection channel 3. As a consequence, the cells (or beads) 8, 9 are prevented from adhering to the side walls 14, 15 of the detection channel 3. By restricting the flow 2 of sample fluid to the detection channel's center portion 13, plugging of the detection channel, which has frequently occurred in prior art detection cells, can be avoided. Furthermore, the side walls 14, 15 may be coated with an anti-adhesive coating like e.g. PEG (polyethylene glycol). [0042] Fig. 1 C shows a cross section of the detection channel 3. The sheath flow 5 flows between the side wall 14 and the flow 2 of sample fluid, whereas the sheath flow 7 is located between the flow 2 of sample fluid and the side wall 15. Thus, a two-sided squeezing of the flow 2 of sample fluid is accomplished. The flow 2 of sample fluid is confined to the detection channel's center region. As a consequence, the cell 9 is prevented from getting too close to any of the side walls 14, 15. In case the sample fluid comprises cells, the detection channel 3 should have about twice the size of a cell's diameter. For cells having a diameter of about 10μm, the detection channel's width might e.g. be 20 to 25 μm.

[0043] The detection cell shown in Figs. 1A, 1B and 1C might e.g. be used for determining an electric, an electromagnetic or an optical property of the sample fluid. If the sample fluid comprises cells or beads, the detection cell might be used for determining an electric, electromagnetic or optical property of the cells or beads. For this purpose, a stimulus signal is applied to the sample fluid, and in response to this stimulus signal, a response signal is detected and evaluated.

[0044] In the embodiments shown in Figs. 1A, 1B and 1C, the detection cell comprises a transmitter electrode 16 and a receiver electrode 17. Via the transmitter electrode 16, an electrical stimulus signal is coupled to the sample fluid. The electrical stimulus signal might either be a DC signal or an AC signal. The receiver electrode 17 is adapted for receiving, in response to the stimulus signal, an electrical response signal. If an AC signal is applied to the transmitter electrode 16, a related AC response signal will be received by the receiver electrode 17. In this case, it is possible to implement both the transmitter electrode 16 and the receiver electrode 17 as contactless electrodes, because AC signals may be capacitively coupled to the sample fluid. Coupling from and to the electrodes 16, 17 might e.g. take place at a location where the sample flow is not (yet) confined, thus enhancing coupling efficiency. Alternatively, if a DC signal is applied to the transmitter electrode 16, a DC response signal will be received by the receiver electrode 17. In case of DC signals, both the transmitter electrode 16 and the receiver electrode 17 are in direct contact with the sample fluid. Again, coupling takes place at a location where the sample flow is not confined, which ensures good contact from electrodes on the channels' surface to the sample liquid.

[0045] The received AC or DC response signal 18 is forwarded to a determination unit 19 for further analysis. In the determination unit 19, one or more electrical properties of the sample fluid, like e.g. conductivity, complex conductivity, impedance, resistance, reactance, relative permittivity, etc. might be determined.

[0046] In case the sample fluid comprises cells (or beads) 8, 9, the detection cell of Figs. 1A, 1B and 1C can be used for counting or analyzing these cells or beads, because each time a cell 9 enters the detection channel 3, a change of the measured electrical property can be detected. The determination unit 19 might e.g. count the number of cells passing through the detection channel 3 per unit time, and derive the cells' concentration therefrom.

[0047] It is advantageous if the conductivity of the sample fluid exceeds the conductivity of the sheath fluid. For example, the sample fluid might comprise a higher concentration of charged ions than the sheath fluid. Thus, the sheath flows 5, 7 substantially act as an electrical insulation sheathing the sample fluid, whereas the sample fluid may be seen as a "liquid wire" adapted for contacting a cell (or bead) 9. The response to an applied AC or DC stimulus signal can be understood using an electrical model, whereby the upper portion 20 of the sample flow, the cell 9, and the lower portion 21 of the sample flow are considered as a series connection of three electric components. Due to the presence of the sheath flows 5, 7, the flow 2 of sample fluid is restricted to the center portion 13 of the detection channel 3. Hence, the cells 8, 9 are prevented from attaching to the side walls 14, 15 of the detection channel 3.

[0048] Confining the sample fluid to the center portion 13 of the detection channel is also advantageous in view of the measurement accuracy of the set-up. In prior art solutions, the AC or DC current flowing on the right-hand and left-hand side of the cell 9 has provided a major contribution to the total AC or DC current. In the detection cell shown in Figs. 1A, 1B and 1C, the contribution to the AC or DC current flowing in parallel to the cell 9 is considerably reduced. The measured AC or DC current substantially represents the current flowing through the cell 9 itself. In this regard, the sheath flows 5, 7 help to improve the sensitivity of the measurement set-up. In fact, the upper portion 20 and the lower portion 21 of the flow 2 of sample fluid can be considered as contact wires adapted for properly contacting the cell 9.

[0049] Thus, the sheath flows 5, 7 are helpful for determining the electrical properties of the cell 9. In addition to counting the cells 8, 9, the detection cell of Figs. 1A, 1B and 1C permits measuring the cells' electrical properties. For example, it is possible to distinguish different types of cells (or beads) according to their electrical behavior. For example, an AC stimulus signal comprising different AC frequency components might be coupled to the transmitter electrode 16. If the frequency of a certain AC frequency component corresponds to an activation energy of the cell 9, this frequency component will be absorbed by the cell 9. For example, a certain AC frequency component might correspond to an activation energy for rotatory movements of the cell 9. Therefore, one or several frequency components might be absorbed by the cell 9, while others might not experience any attenuation. By comparing the frequency spectrum of the received AC response signal with the stimulus signal's frequency spectrum, it is possible to derive characteristic properties of the cells, and to distinguish different types of cells.

[0050] Fig. 2 shows another embodiment of a detection cell according to the present invention. Via a first inlet 22, a flow 23 of sample fluid is supplied to the detection cell, and via the second inlets 24, 25, sheath flows 26, 27 are supplied. In contrast to the embodiment shown in Figs. 1 A, 1 B and 1 C, the detection cell of Fig. 2 comprises one common outlet 28 for draining off both the sample fluid and the sheath fluid. The detection cell further comprises a transmitter electrode 29 and a receiver electrode 30. Over the length of the detection channel 31 , the flow 23 of sample fluid and the sheath flows 26, 27 flow in parallel to one another. Due to lateral diffusion, the two different liquids start to mix. The extent of lateral diffusion mainly depends upon the time required by the two fluids for traversing the detection channel 31. By keeping the detection channel's length rather small (e.g. up to several hundred micrometers), it can be made sure that phase separation between the two liquids is substantially maintained. Downstream of the detection channel 31 , the two liquids might start to mix more significantly, but this is of no importance for the proposed detection technique. [0051] Use of a detection cell according to embodiments of the present invention is not restricted to detecting electric or electromagnetic properties of the sample fluid. Fig. 3 shows a perspective view of a detection cell that is adapted for determining an optical property of a sample fluid. This detection cell comprises a first inlet 32 for supplying the sample fluid 33, and second inlets 34 for supplying a sheath fluid 35. The sample fluid is drawn off via a first outlet 36, whereas the sheath fluid is drawn off via second outlets 37. Due to the two-sided squeezing, the flow of sample fluid 33 is confined to a center portion 38 of the detection cell. A light source 39, preferably a laser source, provides an incident beam of light 40 that is focussed onto the center portion 38 of the detection cell. The measurement set-up may further comprise a first detection unit 41 adapted for measuring the intensity of a forward radiated or scattered signal 42. From the intensity of the forward radiated signal 42, absorption of the sample fluid 33 may be derived. Alternatively or additionally, the measurement set-up may comprise a second detection unit 43 arranged at an angle of 90° relative to the incident beam of light 40. The second detection unit 43 is adapted for determining the intensity of a side-scattered signal 44. The measurement set-up of Fig.3 can also be used for detecting fluorescence intensity of fluorescent labeled compounds contained in the sample fluid 33. For example, the fluorescence measurement set-up of Fig. 3 can be used for detecting fluorescence intensity of cells (or beads) 45 contained in the sample fluid 33. The incident beam of light 40 is used for exciting the fluorescent labels of the labeled compounds. The re-emitted fluorescence light, which usually has a larger wavelength than the incident light, is either detected by the first detection unit 41 , or by the second detection unit 43, or by both of them.

[0052] The optical detection technique shown in Fig. 3 may be combined with the electrical detection shown in Figs. 1 A, 1 B and 1 C. For this purpose, the detection cell of Fig.3 may additionally comprise a transmitter electrode 46 for coupling an electrical stimulus signal to the sample fluid 33, and a receiver electrode 47 for receiving an electrical response signal. However, the optical detection cell of Fig. 3 does not have to comprise an additional electrical detection mechanism, and for this reason, the transmitter electrode 46 and the receiver electrode 47 are illustrated with dashed lines.

In case the sample fluid 33 comprises fluorescent-labeled cells (or beads) 45, the electrical response signal provided by the receiver electrode 47 can be used for triggering the detection of fluorescence intensity. For example, whenever a cell (or bead) 45 enters the detection channel, a corresponding change of the sample fluid's conductivity is detected. This change of conductivity indicates that there is a cell 45 passing through the detection channel. Whenever a change of conductivity of this kind is encountered, fluorescence intensity of the respective cell will be evaluated. Furthermore, the electrical detection might be used for detecting doublets of cells passing through the detection channel. Whenever the response signal indicates that two cells contribute simultaneously to the detected fluorescence intensity, the corresponding measurement value will be sorted out.

[0053] The measurement set-up of Fig. 3 might e.g. be used for tracking fluorescence intensity of fluorescent labeled cell DNA during cell cleavage. Just before a cell is segmented into two daughter cells, the amount of DNA contained in the cell is significantly increased (doubled). This increase of the amount of DNA leads to a corresponding increase of the detected fluorescence intensity. Accordingly, by tracking fluorescence intensity of cells passing through the detection channel, the process of cell cleavage can be monitored.

[0054] The measurement set-up of Fig. 3 can further be used for observing fluorescence intensity of functionalized polymeric beads, such as the functionalized polymeric beads 48 shown in Fig. 4. The polymeric beads 48 have been chemically modified and comprise a number of functional groups 49 adapted for reacting with certain chemical compounds, or with a class of chemical compounds. Using beads of the kind shown in Fig. 4, the presence or absence of certain chemical compounds might affect at least one of the optical or electrical properties of the beads 48. A measurement set-up of the kind shown in Fig. 3 allows tracking these optical and/or electric properties.

[0055] Fig. 5 gives an overview of the parameters that affect the geometry of the flows in the detection cell. Sample fluid at an atmospheric pressure p is supplied to a first inlet 50. Similarly, sheath fluids at an atmospheric pressure p is provided to second inlets 51 , 52. In order to draw off the sample fluid, a sub-atmospheric pressure Pi is applied to a first outlet 53. A flow 54 of sample fluid is established, which is driven mainly by the pressure difference (p-pi) applied across the length I₁ of the sample flow path. In order to withdraw the sheath fluid, a sub-atmospheric pressure p₂ is applied to the outlets 55, 56. As a consequence, sheath flows 57, 58 are established. The sheath flows 57, 58 are driven mainly by the pressure difference (p-p₂) applied across the length I₂ of the respective sheathing flow paths.

[0056] The sub-atmospheric pressures pi and p₂, which might e.g. be in the range between 10 mbar and 400 mbar, can be applied by connecting the outlets 53, 55, 56 to one or more vacuum pumps. For example, for generating the required sub-atmospheric pressures pi, p₂, a waterjet air pump or a peristaltic pump might be used.

[0057] The volumetric flow, which is defined as a volume per unit time, depends on a conduit's hydraulic resistance R, and on a pressure difference Δp applied across the conduit. The volumetric flow can be approximated as follows:

[0058] J = ^ = ^V R η l

[0059] with η denoting the fluid's viscosity, with A denoting the conduit's cross section, and with I being the length of the conduit. However, it has to be kept in mind that this formula only provides an approximation, because in a radial direction, different pressures inside the flow cell level out.

[0060] Because of the presence of the sheath flows, the flow of sample fluid is confined to a relatively small cross section A₂ , and the sample fluid's linear speed is increased. The cross section A₂ is considerably smaller than the detection channel's total cross section A. In fact, in the detection channel's center portion, the sample flow's increased velocity v₂ is mainly determined by the degree of squeezing. Due to the equation of continuity

[0061] V₁ A₁ = V₂ A₂

[0062] the ratio of the velocities V₁ and V₂ is substantially equal to the ratio of the cross sections A₁ and A₂, with A₁ denoting the cross section at the inlet 50, and with A₂ denoting the cross section of the sample flow 54 in the detection channel's center portion. As a consequence, the smaller the effective cross section A2 of the sample flow 54 is, the higher the corresponding local velocity V₂ will be. The detection channel's total cross section A is another parameter that can be used for sizing the respective flows in the detection cell.

[0063] Fig.6 shows a technical implementation for realizing a two-sided squeezing of a flow of sample fluid. The flow 59 of sample fluid is conveyed through a tubing 60. Via the two conduits 61 , 62, respective flows 63, 64 of sheath fluid are supplied. As a consequence, in region 65, a two-sided squeezing of the flow 66 of sample fluid is accomplished. By means of the two sheath flows 67, 68, the flow 66 is confined to a center portion of the tubing 60. The outlet 69 might be a common outlet adapted for drawing off both the sample fluid and the sheath fluid. Alternatively, the detection cell might further comprise additional outlets adapted for drawing off the sheath fluid. By reducing the relative size of the flow 59 of sample fluid, the two sheath streams may mate at top and bottom, and if this happens, an all-sided squeezing will be accomplished.

[0064] A detection cell according to embodiments of the present invention can be implemented as a microfluidic chip device. The fluid passageways of the detection cell shown in Fig. 6 can be realized using microstructuring techniques. For example, the detection cell might be implemented as a dual-layer structure comprising a lower layer 70 and an upper layer 71. Microstructuring techniques such as e.g. etching, hot embossing, laser ablation or micromolding might be used for processing the layers' surfaces.

[0065] Fig.7 shows an alternative embodiment of a detection cell that provides for an all-sided squeezing of the sample flow. A flow 72 of sample fluid is supplied to a conduit 73. Via four conduits 74, 75, 76, 77, respective sheath flows 78, 79, 80, 81 are supplied. Hence, in the detection channel 82, an all-sided squeezing of the flow 83 of sample fluid is accomplished. The embodiment of Fig. 7 can be implemented as a multilayer structure comprising layers 84, 85. However, for accommodating fluid passageways adapted for supplying sheath fluid to the conduits 75, 77, respectively, it will be necessary to add additional layers.

Claims

1. A fluid cell comprising

a first inlet (1) adapted for providing a sample fluid to the fluid cell;

a second inlet (4, 6) adapted for providing a sheath fluid to the fluid cell;

one or several outlets (10, 11, 12);

with a sample flow (2) of sample fluid being established between the first inlet (1 ) and one of the one or several outlets (10, 11 , 12), and with a sheath flow (5, 7) of sheath fluid being established between the second inlet (4, 6) and at least one of the one or several outlets (10, 11 , 12), said sheath flow (5, 7) being adapted for sheathing the sample flow (2), at least in a part of the fluid cell;

a receiver adapted for receiving a response signal in response to a stimulus signal coupled to the fluid cell.

2. The fluid cell of claim 1, wherein both the first inlet and the second inlet are located in the area of the fluid cell's first end.

3. The fluid cell of claim 2, wherein the one or several outlets are located in the area of the fluid cell's second end opposite the first end.

4. The fluid cell of claim 1 or any one of the above claims, wherein the stimulus signal is either of an electrical signal, electromagnetic signal, or optical signal.

5. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell comprises a first electrode located in the area of the fluid cell's first end or in the area of the fluid cell's second end, said first electrode being adapted for coupling a stimulus signal to the fluid cell.

6. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell comprises a second electrode located in the area of the fluid cell's second end or in the area of the fluid cell's first end, said second electrode being adapted for receiving a response signal in response to a stimulus signal coupled to the fluid cell.

7. The fluid cell of claim 1 or any one of the above claims, wherein, within the fluid cell, the sample fluid and the sheath fluid are in direct contact with one another.

8. The fluid cell of claim 1 or any one of the above claims, further comprising at least one of the following features:

the fluid cell comprises a determination unit adapted for deriving, from the response signal, an electrical property of the sample fluid;

the stimulus signal is an AC signal, and the response signal is an AC response signal;

the electrical property is at least one of: conductivity, complex conductivity, impedance, resistance, reactance, relative permittivity;

the conductivity of the sample fluid is higher than the conductivity of the sheath fluid.

9. The fluid cell of claim 1 or any one of the above claims, wherein the length of the fluid cell is chosen as to ensure that the sample flow and the sheath flow substantially do not mix.

10. The fluid cell of claim 1 or any one of the above claims, wherein the length of the fluid cell is chosen as to ensure that the sample flow and the sheath flow remain substantially phase separated over the length of the fluid cell.

11. The fluid cell of claim 1 or any one of the above claims, further comprising one of the following features:

the length of the fluid cell ranges from 1 μm to 1800 μm;

the length of the fluid cell ranges from 10 μm to 300 μm.

12. The fluid cell of claim 1 or any one of the above claims, wherein at least one of the first electrode and the second electrode is in direct contact with the sample fluid.

13. The fluid cell of claim 1 or any one of the above claims, wherein the first electrode is adapted for capacitively coupling an AC signal to the fluid cell.

14. The fluid cell of claim 1 or any one of the above claims, wherein the second electrode is adapted for capacitively receiving an AC response signal in response to an AC signal that has been coupled to the detection cell.

15. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell comprises a common outlet adapted for draining off both the sample fluid and the sheath fluid.

16. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell comprises a first outlet adapted for draining off primarily the sample fluid and at least one second outlet adapted for draining off primarily the sheath fluid.

17. The fluid cell of claim 1 or any one of the above claims, further comprising one of the following features:

one single stream of sheath fluid located on one side of the sample flow confines the sample flow in a way that a single-sided squeezing of the sample flow is accomplished;

two streams of sheath fluid located on the right side and on the left side of the sample flow confine the sample flow in a way that a two-sided squeezing of the sample flow is accomplished;

the sheath flow of sheath fluid surrounds the sample flow in a way that an all- sided squeezing of the sample flow is accomplished.

18. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell is adapted for at least one of counting and analyzing cells, beads or a mixture of both contained in the sample fluid.

19. The fluid cell of claim 1 or any one of the above claims, wherein the sample fluid acts as a liquid wire adapted for contacting a cell or beads while those are present in the fluid cell.

20. The fluid cell of claim 1 or any one of the above claims, wherein an electrical property is modulated by the presence or absence of cells in the fluid cell.

21. The fluid cell of claim 1 or any one of the above claims, wherein the determination unit is adapted for detecting changes of the electrical property that indicate that a cell or bead passes through the fluid cell.

22. The fluid cell of claim 1 or any one of the above claims, wherein the determination unit is adapted for spectrally analyzing the response signal.

23. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell's inner surface is coated with a coating in order to reduce adhesion between the cells and the fluid cell's inner surface.

24. The fluid cell of claim 1 or any one of the above claims, wherein the fluid cell is substantially twice as wide as a cell's diameter.

25. The fluid cell of claim 1 or any one of the above claims, further comprising a light source and an optical detection unit.

26. The fluid cell of claim 1 or any one of the above claims, further comprising one of the following features:

a fluorescence detection unit is adapted for evaluating fluorescence intensity whenever the electrical property indicates that a cell or bead passes through the detection cell;

a determination unit is adapted for evaluating the response signal whenever the fluorescence detection unit indicates that a cell or bead passes through the detection cell.

27. The fluid cell of claim 1 or any one of the above claims, further comprising at least one of the following features: the fluid cell is implemented as a microfluidic chip device;

the fluid cell is implemented using microstructuring technologies such as laser ablation, hot embossing, etching, micromolding;

the fluid cell is implemented as a multilayer structure comprising two or more layers.

28. A fluid separation system comprising one or more of

a fluid delivery system,

a separation device for separating components of the fluid delivered by the fluid delivery system, and

a fluid cell of claim 1 or any one of the above claims for at least one of analyzing and detecting components of the fluid separated by the separation device.

29. The fluid separation system of claim 28, wherein the fluid separation system is or comprises at least one of:

a chromatographic system,

a high performance fluid chromatographic system,

a high performance liquid chromatography arrangement comprising a chip and a mass spectrograph,

a high throughput liquid chromatography / mass spectroscopy system,

a purification system,

a micro fraction collection / spotting system,

a system adapted for identifying proteins,

a system comprising a gel permeation chromatography / size exclusion chromatography column, a nanoflow liquid chromatography system, or

a multidimensional liquid chromatography system adapted for separation of protein digests.

30. A method comprising the steps of:

providing a sample fluid via a first inlet (1 ) to a fluid cell;

providing a sheath fluid via a second inlet (4, 6) to the fluid cell;

draining off the sample fluid and the sheath fluid via one or several outlets (10, 11 , 12);

with a sample flow (2) being established between the first inlet (1 ) and one of the one or several outlets (10, 11, 12), and with a sheath flow (5, 7) of sheath fluid being established between the second inlet (4, 6) and at least one of the one or several outlets (10, 11, 12), said sheath flow (5, 7) being adapted for sheathing the sample flow (2), at least in a part of the fluid cell;

coupling a stimulus signal to the fluid cell and receiving a response signal in response to the stimulus signal.

31. The method of claim 30, wherein within the fluid cell, the sample fluid and the sheath fluid are in direct contact with one another.

32. The method of claim 30 or any one of the above claims, further comprising a step of deriving, from the response signal, a property of the sample fluid.

33. The method of claim 30 or any one of the above claims, further comprising at least one of the following steps:

counting cells or beads contained in the sample fluid;

analyzing cells or beads contained in the sample fluid;

correlating an electrical property of the sample fluid with any kind of secondary parameter derived from the cells or beads.

34. The method of claim 30 or any one of the above claims, further comprising a step of detecting changes of the electrical property that indicate that a cell passes through the fluid cell.

35. The method of claim 30 or any one of the above claims, further comprising a step of evaluating fluorescence intensity of the sample fluid in addition to determining an electrical property of the sample fluid.

36. The method of claim 30 or any one of the above claims, further comprising a step of evaluating fluorescence intensity whenever the electrical property indicates that a cell passes through the fluid cell.

37. The method of claim 30 or any one of the above claims, further comprising a step of evaluating the response signal whenever fluorescence intensity indicates that a cell passes through the fluid cell.

38. A software program or product, preferably stored on a data carrier, for controlling the steps of claim 30 or any one of the above claims, when run on a data processing system such as a computer.