The present teachings generally relate to the processing of sample fluids containing one or more analytes of interest, and more particularly, to methods and devices for separating and/or purifying components of a sample fluid using electric and hydrodynamic forces.
Purifying biological materials such as proteins, DNAs, lipids, and metabolites in physiological conditions is essential in understanding life processes. The purification technologies currently available, such as two-dimensional gel electrophoresis, analytical chromatography and capillary electrophoresis, usually require denaturation of biomolecules or are incapable of processing a reasonable amount of material. While capillary electrophoresis (CE), which utilizes the differential mobility of charged species in an applied electric field, can be used to separate species that cannot be reliably resolved by other methods, fraction collection of CE generally produces small amounts of purified sample components due to limitations associated with the physical dimensions of the capillary columns. More recently, two-dimensional electro-fluid-dynamic (2-D EFD) devices have utilized hydrodynamic pressure, as well as an electric field, to drive analyte and fluid migration through two-dimensional channel networks (as opposed to the one-dimensional columns of CE). 2-D EFD devices have been shown to continuously purify multiple components from complex samples into different channels, each containing a substantially pure compound, as described for example in “Reverse of Mixing Process with a Two-Dimensional Electro-Fluid-Dynamic Device,” Anal. Chem., 82:2182-2185 (2010), and “Potential of Two-Dimensional Electro-Fluid-Dynami Devices for Continuous Purification of Multiple Components from Complex Sample,” Anal. Chem., 83:8208-8214 (2011), which list as authors the present inventors et al. and are incorporated by reference in their entireties. In these exemplary 2-D EFD devices, analytes are driven through the fluid by non-discriminative forces (e.g., pressure or electroosmosis) and by discriminative forces (e.g., from the applied electric field). These forces exist simultaneously and produce a net migration of analytes determined by the sum of the velocity vectors. For example, in conventional 2D-EFD devices, a positive electric potential is applied at a sample vial to drive charged analytes within the sample into and against a flow of an electrolytic fluid in a separation stream, of which the pressure-induced velocity can be precisely controlled so as to manipulate the migration of the various analytes in the same direction as or against the bulk flow of fluid in the separation channel due to the electrophoretic mobility of the particular analyte.
There remains a need for improved devices for performing continuous chemical purification and/or separation.
In some aspects, the present teachings provide methods and systems for simultaneously obtaining multiple fractions having simpler compositions, including pure biological compounds solutions from a complex mixture, and can enable the complete processing of a whole sample, without the need for pre-fractionation. As described above, known 2-D EFD utilize a fluid flow in a main separation channel into which one or more analyte can be injected utilizing an electric potential applied at a sample vial to drive charged analytes within the sample into a micro scale channel for separation. In accordance with various aspects of the present teachings, systems and methods in accordance with the present teachings utilize a pressure-driven bulk fluid flow to deliver a sample fluid into the separation stream without discriminating individual analytes based on their charge status, and can provide faster sample processing, resistance to unstable electroosmotic flow, and avoid buffer depletion that is common in known 2D-EFD device. Further, the continuous nature allows the complete processing of the mixture that is constantly introduced. The techniques disclosed herein can also be combined with other separation, reaction, and detection techniques to further characterize the purified fractions or compounds to determine their molecular composition, structure and biological and chemical activities. In various embodiments, a sample channel can comprise a channel network. In various embodiments, a separation channel can comprise a channel network. In various embodiments, a collection channel can comprise a channel network.
In accordance with one aspect, certain embodiments of the applicant's teaching relate to a microfluidic device for separating components of a fluid sample comprising one of a sample channel and channel network extending between an inlet end and an outlet junction, the inlet end configured to receive a fluid sample containing one or more analytes to be delivered to the outlet junction, wherein a fluid pressure of the fluid sample at the inlet end of one of the sample channel and channel network is greater than a fluid pressure at the outlet junction. The device also includes one of a separation channel and channel network in fluid communication with the one of the sample channel and channel network, one of the separation channel and channel network extending from an inlet end to said outlet junction, the inlet end of one of the separation channel and channel network configured to receive a counter-flow fluid, wherein a fluid pressure of the counter-flow fluid at the inlet end of one of the separation channel and channel network is greater than said fluid pressure at the outlet junction. A first collection channel or at least one of a collection channel and channel network is in fluid communication with one of the separation channel and channel network and one of the sample channel and channel network at the outlet junction and a plurality of electrodes generate an electric field within one of the separation channel and channel network and the first collection channel or at least one of the collection channel and channel network. In some aspects, the device can include a sample pump (e.g., a syringe pump) fluidically coupled to the inlet end of one of the sample channel and channel network for pumping the fluid sample from the inlet end of one of the sample channel and channel network to the outlet junction. In various aspects, the electric field and a hydrodynamic force on the one or more analytes at the outlet junction preferentially drive the one or more analytes into one of said separation channel or channel network and said first collection channel or said at least one of the collection channel or channel network.
In various aspects, the microfluidic device can further comprise a counter-flow pump (e.g., a syringe pump) fluidically coupled to the inlet end of one of the separation channel and channel network for pumping the counter-flow fluid from the inlet end of one of the separation channel and channel network to the outlet junction.
In various aspects, one of the sample channel and channel network can comprise a substantially electric field-free region. For example, the inlet end of one of the sample channel and channel network can be electrically floated. In some aspects, the plurality of electrodes are arranged so as to generate a substantially electric field-free region in one of the sample channel and channel network. For example, in related aspects, none of the plurality of electrodes are disposed adjacent the inlet end of one of the sample channel and channel network.
According to various aspects of the present teachings, at least one of the fluid pressure at the inlet end of one of the sample channel and channel network, the fluid pressure at the inlet end of one of the separation channel and channel network, and the electric field generated by the plurality of electrodes can be adjustable such that a first species of analyte is preferentially driven into said first collection channel or one of said at least one collection channel and channel network and a second species of analyte is preferentially driven into one of said separation channel and channel network. In some aspects, the fluid flow velocity through the sample channel, the separation channel, and/or the collection channel can be controlled utilizing one or more pumps.
In some aspects, one of the first collection channel and channel network extends from the outlet junction to a first fluid reservoir, with one of the plurality of electrodes being in contact with fluid in the first fluid reservoir. For example, one of the first collection channel and channel network can extend from the outlet junction to a first fluid reservoir and the device can further comprise one of a second collection channel and channel network extending from one of the separation channel and channel network at a second fluid separation junction spaced a distance apart from the outlet junction, one of the second collection channel and channel network defining a fluid flow pathway between the second fluid separation junction and a second fluid reservoir. In related aspects, one of the plurality of electrodes can be in contact with fluid in the first fluid reservoir and one of the plurality of electrodes can be in contact with fluid in the second fluid reservoir. In some aspects, one of a third collection channel and channel network extending from one of the separation channel and channel network at a third fluid separation junction spaced a distance apart from the outlet junction and the second fluid separation junction can also be provided, one of the third collection channel and channel network defining a fluid flow pathway between the third fluid separation junction and a third fluid reservoir. By way of example, the second fluid separation junction can be disposed between the outlet junction and the third fluid separation junction. In some aspects, a first electrode of the plurality of electrodes can be in contact with fluid in the first fluid reservoir, a second electrode of the plurality of electrodes can be in contact with fluid in the second fluid reservoir, and a third electrode of the plurality of electrodes can be in contact with fluid in the third fluid reservoir. In such aspects, at least one of the fluid pressure at the inlet end of one of the sample channel and channel network, the fluid pressure at the inlet end of one of the separation channel and channel network, and the electric field generated by the plurality of electrodes can be adjustable such that a first species of analyte is preferentially driven into one of said first collection channel and channel network and a second and third species of analyte is preferentially driven into one of said separation channel and channel network. Additionally in some aspects, at least one of the fluid pressure at the inlet end of one of the sample channel and channel network, the fluid pressure at the inlet end of one of the separation channel and channel network, and the electric field generated by the plurality of electrodes can be adjustable such that the second species of analyte is preferentially driven into one of the second collection channel and channel network and the third species of analyte is preferentially driven into one of the third collection channel and channel network.
The electrodes can have a variety of configurations and potentials applied thereto. For example, the first and second electrodes of the at least three electrodes can be equipotential, and wherein at least one of an average cross-sectional area and a channel length of one of the collection channels and channel network differ from one another. In related aspects, a single power source can be electrically coupled to the first and second electrodes for applying an electric potential thereto. Alternatively, an electric potential applied to the first and second electrodes can differ from one another, and wherein at least one of an average cross-sectional area and a channel length of one of the collection channels and channel network are substantially equal. For example, a first and a second power source can be electrically coupled to the first and second electrodes, respectively, for applying an electric potential thereto.
In accordance with one aspect, certain embodiments of the applicant's teaching relate to a microfluidic device for separating components of a fluid sample comprising a pump fluidically coupled to one of the sample channel and channel network for pumping a fluid sample containing one or more analytes from an inlet end of one of the sample channel and channel network to an outlet junction via a substantially electric field-free pathway; one of a separation channel and channel network in fluid communication with one of the sample channel and channel network, one of the separation channel and channel network extending from an inlet end to said outlet junction, the inlet end of one of the separation channel and channel network configured to receive a counter-flow fluid; a first collection channel or at least one of a collection channel and channel network in fluid communication with one of the separation channel and channel network and one of the sample channel and channel network at the outlet junction; and a plurality of electrodes configured to generate an electric field within one of the separation channel and channel network and the first collection channel or at least one of the collection channel and channel network, wherein the electric field and a hydrodynamic force on the one or more analytes at the outlet junction preferentially drive the one or more analytes into one of said separation channel or channel network and said first collection channel or at least one of the collection channel or channel network. In some aspects, at least one of a volumetric flow rate of the fluid sample, a volumetric flow rate of the counter-flow fluid, and the electric field generated by the plurality of electrodes can be adjustable such that a first species of analyte is preferentially driven into said first collection channel or one of said at least one collection channel and channel network and a second species of analyte can be preferentially driven into one of said separation channel and channel network.
In accordance with one aspect, certain embodiments of the applicant's teaching relate to a method of separating fluids comprising: pumping a sample fluid from an inlet end of one of a sample channel and channel network to an outlet junction; pumping a counter-flow fluid from an inlet end of one of a separation channel and channel network to the outlet junction; and generating an electric field in the one of the separation channel and channel network such that one or more analytes in the sample fluid at the outlet junction are preferentially driven into one of said separation channel and channel network and a first collection channel or at least one of a collection channel and channel network in fluid communication with one of the separation channel and channel network and one of the sample channel and channel network at the outlet junction.
In some aspects, the method can also include adjusting at least one of a volumetric flow rate of the fluid sample, a volumetric flow rate of the counter-flow fluid, and the electric field such that a first species of analyte is preferentially driven into said first collection channel or one of a respective collection channel and channel network and a second species of analyte is preferentially driven into one of said separation channel and channel network.
In various aspects, one of the sample channel and channel network can be a substantially electric field-free pathway. In some aspects, the sample fluid can be pumped at a substantially constant volumetric flow rate while adjusting a volumetric flow rate of the counter-flow fluid so as to effect an interaction between a hydrodynamic force and electric field experienced by the one or more analytes at the outlet junction, e.g., so as manipulate along which channel is preferentially driven.
In some aspects, the first collection channel or at least one of the collection channel and channel network can extend from the outlet junction to a first or respective fluid reservoir, a second collection channel or channel network can extend from the separation channel or channel network at a second fluid separation junction spaced a distance apart from the outlet junction (the second collection channel or channel network defining a fluid flow pathway between the second fluid separation junction and a second fluid reservoir), and a third collection channel or channel network can extend from the separation channel or channel network at a third fluid separation junction spaced a distance apart from the outlet junction (the third collection channel or channel network defining a fluid flow pathway between the third fluid separation junction and a second fluid reservoir), wherein a first electrode is in contact with fluid in the first fluid reservoir, a second electrode is in contact with fluid in the second fluid reservoir, and a third electrode is in contact with fluid in the third fluid reservoir. In a related aspect, the method can comprise adjusting at least one of a volumetric flow rate of the fluid sample, a volumetric flow rate of the counter-flow fluid, and an electric potential applied to one of the first, second, and third electrodes such that a first species of analyte is preferentially driven into said first or respective collection channel or channel network and a second and third species of analyte is preferentially driven into said separation channel or channel network.
In related aspects, the method can also include adjusting at least one of the volumetric flow rate of the fluid sample, the volumetric flow rate of the counter-flow fluid, and the electric potential applied to one of the first, second, and third electrodes such that the second species of analyte is preferentially driven into one of a respective collection channel or channel network, i.e., second collection channel or channel network and the third species of analyte is preferentially driven into said respective or third collection channel or channel network.
In some aspects, at least one of an average cross-sectional area and a channel length of the first and second collection channels or one of the collection channel and channel network can differ from one another, and the method can further comprise maintaining the potential applied to the first and second electrodes substantially equal. For example, a single power source can be used to generate an electric potential at the first and second electrodes.
Alternatively, at least one of an average cross-sectional area and a channel length of the first and second collection channels or one of the collection channels and channel network can be substantially equal to one another, the method further comprising applying an electric potential of different magnitudes to the first and second electrodes. For example, a first and a second power source can be electrically coupled to the first and second electrodes, respectively, for applying an electric potential thereto.
In accordance with one aspect, certain embodiments of the applicant's teaching relate to a microfluidic device for separating components of a fluid sample, comprising: one of a sample channel and channel network extending from an inlet end fluidically coupled to a reservoir of a fluid sample to a first intersection junction, the fluid sample containing one or more analytes; one of a separation channel and channel network in fluid communication with one of the sample channel and channel network, one of the separation channel and channel network extending from an inlet end to said first intersection junction, the inlet end of one of the separation channel and channel network configured to receive a counter-flow fluid; a first collection channel or at least one of a collection channel and channel network in fluid communication with one of the separation channel and channel network at the first intersection junction, one of the first or respective collection channel and channel network extending from the first intersection junction to a first or respective collection reservoir in contact with a first or respective electrode to which a first or respective electric potential can be applied; and a second collection channel or channel network in fluid communication with the separation channel or channel network at a second intersection junction spaces apart from the first intersection junction, the second collection channel or channel network extending from the second intersection junction to a second or respective collection reservoir in contact with a second electrode to which a second electric potential can be applied. The electrodes are configured to generate an electric field within one of the separation channel and channel network and the first and second collection channels or at least one of the respective collection channel and channel network. At least one of an average cross-sectional area and channel length of the first and second collection channels or one of the collection channels or channel network differ from one another. In some aspects, the first and second electrodes are equipotential, and the potential can be applied by a single power source. In some aspects, the device can include a sample electrode for electrokinetically driving the analytes within the sample fluid from the sample inlet to the first intersection point.
These and other features of the applicant's teachings are set forth herein and in the appendix attached hereto, which is hereby incorporated by reference in its entirety.
The skilled person in the art will understand that the drawings, described below, are for illustration purposes only. The drawings are not intended to limit the scope of the applicant's teachings in any way.
It will be appreciated that for clarity, the following discussion will explicate various aspects of embodiments of the applicant's teachings, while omitting certain specific details wherever convenient or appropriate to do so. For example, discussion of like or analogous features in alternative embodiments may be somewhat abbreviated. Well-known ideas or concepts may also for brevity not be discussed in any great detail. The skilled person will recognize that some embodiments of the applicant's teachings may not require certain of the specifically described details in every implementation, which are set forth herein only to provide a thorough understanding of the embodiments. Similarly it will be apparent that the described embodiments may be susceptible to alteration or variation according to common general knowledge without departing from the scope of the disclosure. The following detailed description of embodiments is not to be regarded as limiting the scope of the applicant's teachings in any manner. On the contrary, the applicant's teachings encompass various alternatives, modifications, and equivalents, as will be appreciated by those of skill in the art. The features illustrated or described in connection with one exemplary embodiment may be combined with the features of other embodiments. Such modifications and variations are intended to be included within the scope of the applicants' disclosure. Various terms are used herein consistent with their customary meanings in the art. The term “about” as used herein indicates a variation of less than 10%, or less than 5%, or less than 2%.
The present teachings generally relate to the processing of sample fluids containing one or more analytes of interest, and more particularly, to methods and devices for separating and/or purifying components of a sample fluid using electric and hydrodynamic forces. Though the fluid processing systems and methods are generally described herein as applied to microfluidics, it will be appreciated in light of the present teachings that the fluid processing systems may process any fluid volume suitable for use in embodiments described herein. As discussed above, Y-shaped and multiple-branched shaped 2-D EFD devices have been used to separate and/or purify one or more analytes from a mixture. In known devices, an electric field in the sample channel or channel network is utilized to overcome the effect of back pressure to deliver the analytes to be purified into the separation stream (generally referred to herein as electrokinetic injection). However, as discussed in detail below, systems and methods in accordance with various aspects of the present teachings instead utilize hydrodynamic pressure (e.g., using a pump) to drive the sample liquid from the sample inlet to the separation stream, and can, in some aspects, provide improved control of the movement of the analytes, improved processing times, and decreased buffer depletion.
With reference now to
The inlet end (A) of the sample channel (AC) can have a variety of configurations but is generally configured to receive thereat a fluid sample containing one or more analytes to be separated and/or purified by the microfluidic device 100, as indicated by the upper arrow in
The inlet end (F) of the separation channel (CD) can also have a variety of configurations but is generally configured to receive thereat a fluid delivered under pressure to the intersection point (C), as indicated by the lower arrow in
As shown in
In some aspects, the plurality of the electrodes of the microfluidic device 100 can be configured such that the sample channel (AC) is generally a field-free region (e.g., analytes are driven to the intersection point (C) through non-discriminative forces such as a positive pressure differential between the sample inlet (A) and the outlet junction (C) and are not subject to a substantial electric forces). For example, as shown in
It will be appreciated that devices in accordance with the present teachings can be manufactured using any of a plastic, polymer such as PDMS (e.g., Sylard184, Dow Corning, Midland, Mich.), glass, or any other suitable material(s) into which the channels described herein can be formed. By way of non-limiting example, the substrate can comprise soda lime glass (Nanofilm, Westlake Village, Calif.) within which channels are formed using known photolithographic patterning and wet chemical etching methods.
With reference now to
Like the exemplary device of
It should be appreciated, however, in light of the present teachings that the channels for processing the fluids can have a variety of dimensions and/or cross-sectional shapes. Though the calculations presented below regarding exemplary fluid and electric fields present in the various channels of the exemplary devices are presented with regard to symmetric channels (having identical cross sections) or of a fixed ratio of cross-sections, theoretical calculations can likewise be determined in light of the present teachings for channels of any cross-sectional area. For example, though the theoretical calculations and experimental results demonstrate the use of an exemplary symmetrical Y-shaped device 100 in which the channels (AC) and (BC) are symmetrical about the separation channel (CD) and a multiple-branched device 300 in which the width of the main separation channel (CD) is 80 um and the width of the sample channel (AC) and collection channel(s) (BC) is 40 μm (each has a depth of 50 μm), it should be appreciated that these values (both absolute and relative to one another) are merely exemplary and do not necessarily limit the present teachings. For example, the width of the sample channel (AC) and collection channel (BC) could be 50 μm, the width of the main channel (CD 100 μm, and the depth of all PDMS channels could be 200 μm.
Likewise, with reference again to
With reference now to
In the discussions below, the directions of vectors are all along the channel length. Thus, these vectors are expressed as scalars, and the values are defined as positive when the vector direction is toward the intersection point C. The cross-sectional area of channel AC and BC are the same for the exemplary device 100, and the cross-sectional area of channel AC and CD are the same for the exemplary device 300. The cross-sectional area ratio of lateral channels and the main channel is defined as a for both the exemplary Y-shaped EFD device 100 of
The conductivity of the solution in the exemplary devices 100, 300 of
σAC=σBC=σCD (1)
where σ is the conductivity of the solution inside each channel. From Kirchhoff's law, the net current at the intersection is zero.
J
AC
S
AC
+J
AC
S
BC
+J
AC
S
CD=0 (2)
in which J is the current density in each channel and S represents the cross-sectional area of the respective channel. If Ohm's law J=σE is used in eq. (2), it becomes
E
AC
S
AC
+E
AC
S
BC
+E
AC
S
CD=0 (3)
For the exemplary 2-D EFD devices 100, 300 of
Assuming the fluid in the exemplary EFD devices 100, 300 is incompressible, the fluid velocities in the intersecting channels has the following relationship:
vf,ACSAC+vf,BCSBC+vf,CDSCD=0 (5)
Reformatting Eq. (5) according to the cross-sectional area relationships of the channels results in αvf,AC+αvf,BC+vf,CD=0 for the exemplary symmetrical Y-shaped device 100 of
In accordance with the present teachings, the sample fluid (and the one or more analytes therein) is hydrodynamically driven (e.g., pumped via a syringe pump) through the sample channel (AC) to the outlet junction (C) at a fixed net fluid velocity (vinj). Therefore, the hydrodynamic fluid field distribution relationships can be written as and αvinj+αvf,BC+vf,CD=0 and vinj+αvf,BC+vf,CD=0 for the exemplary EFD devices 100, 300, respectively.
The steady state velocity of a charged particle that is moving in a channel can be written as
v=v
ep
+v
eo
+v
p=μepE+μeoE+vp=μepE+vf (6)
where electrophoretic velocity (vep) is discriminative and determined by its electrophoretic mobility (μep), an intrinsic property for a particular analyte. Electroosmotic velocity (veo) and pressure-induced velocity (vp), on the other hand, are non-discriminative and affect all components equally. As illustrated in
Whereas an analyte undergoing electrokinetic injection into a separation stream could have four possible mass transfer pathways according to the various combinations of electric field and pressure, in systems and methods according to the present teachings, the applied pressure delivers the analyte mixture into the device at the velocity vinj such that each analyte can have only three possible mass transfer pathways. For example, in prior electrokinetic injection techniques, the steady-state velocity of the analyte in either the injection channel (AC), the collection channel (BC) and the separation channel (CD) would be in the same direction as the counter-flow when the pressure is high. As such, the analyte would be forced toward the sample inlet A and would not migrate into the sample channel AC or any other channels. Since the positive potential at point A is typically higher than that at point B, as the magnitude of applied pressure is reduced, the steady-state velocity of the analyte could be reversed first in the sample channel (AC), thereby making the analyte migrate through the outlet junction (C) and into the collection channel (BC) (i.e., to the collection vial B). As the pressure is further reduced, analytes at the intersection point (C) can migrate into both the collection channel (BC) and the separation channel (CD). Finally, in the fourth condition, when the counter-pressure in the electrokinetic injection mode is very low, all analytes would migrate along the direction of the electric field, from the sample inlet (A), through the intersection point (C) and enter the separation channel (CD).
In accordance with various aspects of the present teachings, the applied pressure delivers the analyte mixture into the device (e.g., to the outlet junction (C)) at the velocity vinj such that when the counter-pressure is high, the analytes are pushed into the collection channel (BC). As the magnitude of the pressure reduces, the analytes can migrate into either collection channel (BC) or separation channel (CD). When the counter-pressure is very low, all the analyte migrate along the direction of electric field and the analyte at point C follows the migration pathway of A-C-D. The magnitude of fluid velocity in the collection channel (BC) at critical boundary conditions between these three possible mass transfer pathways can be determined to be EBCμep+vinj and EBCμep for the exemplary symmetrical Y-shaped device 100, and and
and EBCμep for the exemplary multiple-branched device 300, depicted in
With reference now to
Whereas in current 2D-EFD electrokinetic injection devices, critical boundary conditions between the four possible mass transfer pathways are dependent on the relative electric field strengths of the particular channel(s) into which the analytes migrate, the above-discussed critical boundary conditions in an exemplary system in accordance with the present teachings demonstrate that differences between −vf,BC values at the critical boundary conditions are independent of the electric field strength, and instead are only determined by the sample injection speed. It will be appreciated that this characteristic of methods and systems in accordance with the present teachings provides a more convenient approach to control the migration behavior of the analyte in the EFD device, as discussed in more detail below.
The critical boundary value (CBV) is defined as the value of −vf,BC at the critical boundary condition (CBC). As discussed above, the fluid velocity in the collection channel (BC) at the CBCs is crucial to manipulating migration behavior of the analyte in the fluid processing systems and methods of the present teachings. For example, by changing selected parameters of the devices 100, 300 (e.g., manipulating a sample or counter pressure syringe pump to control fluid velocity or manipulating applied electric potential to effect the electric field in the collection channel (BC)), it should be appreciated that the CBVs can be easily manipulated into the appropriate value so as to force the components to follow a desired migration pathway. Because it would be impossible to change the electric field strength in only one channel by simply adjusting only one electric potential (as shown in Eq. 3) when utilizing electrokinetic injection without also changing other boundary conditions, CBVs in known devices exhibit complex relationships that may make it difficult to control analyte migration behavior.
In methods and systems in accordance with the present teachings, however, the sample fluid containing the one or more analytes to be separated can be introduced into the EFD device by hydrodynamic pressure (e.g., by pumping the sample fluid through the sample channel), such that the electric field in the collection channel (BC) and separation channel (CD) is only dependent on the difference between the electric potentials applied at points B and E. From the critical boundary conditions illustrated in
for exemplary device 300), where the analyte changes the migration direction in channel CD, can be manipulated with a magnitude of Δvinj for the symmetrical Y-shaped EFD device 100 and
for tne multiple-branched EFD device 300. It should thus be appreciated in light of the present teachings that changing the sample injection speed can provide a convenient approach to adjusting the difference between two the CBVs, while the second CBV (Y) nonetheless remains unchanged.
Likewise, it should also be appreciated in light of the present teachings that adjustments to the electric field in the collection channel (BC) and the separation channel (CD) (e.g., by changing the relative potential of point (B) to point (E)) can also be utilized to control CBV values in systems and methods described herein. Because the difference between the two CBVs (i.e., X relative to Y) is only dependent on the sample injection speed, a change of the electric field strength is effective to move the two critical boundary values with the same magnitude of ΔEBCμep for both the exemplary symmetrical Y-shaped device 100 and the multiple-branched device 300.
The combined utilization of these two approaches can provide a convenient and powerful approach to regulate the absolute and relative positions of the two CBCs. By way of example, the position of the second CBC (Y), at which the steady-state migration velocity of the analyte reverses in the collection channel (BC), can be manipulated by adjusting the electric potential at the point (B), for example, and the position of the first CBC (X) can then be set by controlling the difference between the two CBCs, by way of changing sample injection speed.
Because the CBVs are dependent on the electrophoretic mobility of the analyte, the present teachings provide for the purification and/or separation of one or more species of analytes in the sample fluid based on their distinctive migration pathways at certain electric field and hydrodynamic pressure conditions. As such, analyte species can be preferentially directed into specific collection locations. For example, in order to achieve this continuous chemical purification, the applied electric potential and counter pressure can make the slowest migrating components of the sample fluid follow the pathway of A-C-B to collection vial B, while all of the faster migrating components can exhibit a migration pathway of A-C-D by being preferentially driven into the main separation channel CD at the outlet junction (C) of the sample channel (AC). In order to obtain such a separation between the slow and fast moving sample components in the exemplary symmetrical Y-shaped EFD device 100, the magnitude of the net fluid velocity in collection channel (BC) should be within the range of:
E
BCμep,slow+vinj<−vf,BC<EBCμep,fast (7)
Consequently, as long as the sample injection speed is kept in the range of
v
inj
<E
BC(μep,fast−μep,slow) (8)
the magnitude of the net fluid velocity in the collection channel (BC) can be selected to be within the range indicated in Eq. (7) in order to achieve the continuous chemical purifications. Because of the existence of the maximum injection speed, the minimum sample mixture processing time can be calculated:
in which t is the time required to process the sample mixture with a volume of Vtot.
On the other hand, for the exemplary multiple-branched device 300 in accordance with various aspects of the present teachings, the magnitude of the fluid velocity in the collection channel (BC) during continuous chemical purification occur is as follows:
As such, the requirement of the sample injection speed is:
v
inj
<αE
BC(μep,fast−μep,slow) (11)
and the minimum sample processing time is
With reference now to
Likewise, with reference to
Comparison with Electrokinetic Injection in Continuous Chemical Purification
As discussed above, systems and methods in accordance with various aspects of the present teachings can provide a more convenient approach for controlling the position of critical boundary conditions relative to known devices. Moreover, as demonstrated below for example with reference to the exemplary Y-shaped EFD device, sample processing speed and resistance to fluctuating electroosmotic flow (EOF) can also be increased in systems in accordance with the present teachings to provide improved operability in the continuous chemical purification process.
Because of the continuous nature of the chemical purification provided by the present teachings and known 2D-EFD devices, the amount of an analyte injected into the separation channel during a certain time period should equal the amount of an analyte processed and collected during that time period. It will thus be assumed that the sample processing speed can be described by the injection speed. In accordance with the present teachings, the injection speed for every analyte should be substantially the same (vinj), which as described above in Eq. (8) should be selected to achieve the sample continuous purification as follows: vinj<EBC(μep,fast−μep,slow). In electrokinetic injection however, the speed of delivering components into the EFD device is analyte dependent, as follows:
v
inj
=E
AC(μeo+μep)+vp,AC (13)
For example, if the counter-pressure during electrokinetic injection is relatively high, the analyte remains at the injection point A, and the injection speed is negative (or zero). However, when the counter pressure is reduced and the analyte follows the migration path of A-C-B, the magnitude of the pressure-induced velocity in the injection channel AC is in the range of
and the range of the injection speed of the analyte in this migration situation is:
Similarly, if the analyte has the migration pathway of both A-C-B and A-C-D, the injection speed range is:
If the analyte migrates along the way of A-C-D, the injection speed range is
v
inj>(EAC−EBC)(μeo+μep) (16)
Thus, for the continuous separation of analytes when utilizing electrokinetic injections, the pressure-induced velocity in the sample channel (AC) should be controlled in the range of making the faster migration components have the migration path of A-C-D, while the slower components have the migration path of A-C-B, which is
Accordingly, in electrokinetic injection, the sample processing speed for the mixture is limited by the injection speed of the slower component, which is
Assuming that the electric fields in the collecting channels have the same strength in both the exemplary devices 100, 300 described herein in accordance with the present teachings and those device that instead utilize electrokinetic injection, the difference in the maximum injection speed is:
Because the value of Eq. (19) is positive during the continuous chemical purification process, present teachings that utilize hydrodynamic forces to deliver the sample fluid to the outlet junction can provide faster sample processing speeds (assuming electric field strength in the collection channel (BC) is the same in each of the injection modes). If the electrical voltage at point (B) (not EBC) is kept the same in both modes, an additional electric potential applied at the sample inlet (A) in the electrokinetic injection approach could further decrease the value of EBC, resulting in an even slower sample processing. As such, methods and systems in accordance with the present teachings can provide a faster sample processing speed compared with known electrokinetic sample injection.
Because surface properties of the microfluidic channel wall can change over time during the continuous chemical purification process, the application of a positive electrical potential at the sample inlet as in electrokinetic injections could induce electrolysis of the buffer solution, thereby altering the buffer pH and making the EOF unstable as well, which could additionally effect the sample processing speed and analyte migration velocity.
In methods and systems in accordance with the present teachings, the electric field strength within the injection channel is substantially zero. For example, the current flowing through the injection channel AC could be monitored and adjusted to be zero. The sample injection speed is controlled by manipulating, for example, a pump (e.g., a syringe pump) utilized to deliver the sample fluid through the sample channel (AC) from the inlet end (A) to the outlet junction (C). The range of the injection speed during the continuous chemical purification is 0<vinj<EBC(μep,fast−μep,slow), which is EOF independent. Because the counter-flow can also be controlled in the way of volumetric flow rate by the counter-flow pump (e.g., a second syringe pump), it can be assumed that the net fluid velocity in the main separation channel (CD) also remains the same under different EOF conditions. Accordingly, in channel CD, the net velocity of the component is as follows:
v
CD
=E
CDμep+vf,CD (20)
which is not affected by the EOF value. Because the migration velocity in the sample channel (AC) can be fixed at vinj, due to the effective volumetric flow rate conservation principle, the migration velocity in collection channel (BC) can be unaffected by an unstable EOF.
In electrokinetic injection, however, sample injection speed, described by Eq. (13), can be rearranged as follows because
Due to the electric field strength relationship (αEAC+αEBC+ECD=0), and that EAC>EBC during the purification process in a symmetrical Y-shaped device,
doesn't equal to zero in electrokinetic injection, and thus may be susceptible to an unstable EOF and fluctuation of the sample injection speed.
Moreover, as described in Eq. (17), in the electrokinetic injection mode, the magnitude of the pressure-induced velocity in collection channels should be kept in the range of
During the continuous chemical purification process, the injection speed for the faster moving component (vinj,fast=EAC(μeoμep,fast)+vp,AC) is in the range of
and for the slower migration component, the injection speed range (vinj,slow=EAC(μeo+μep,slow)+vp,AC) is:
Therefore, the possible range for the sample injection speed is also EOF dependent for both faster and slower migration analytes.
Accordingly, the sample injection speed, as well as the range of possible injection speed flow rates, are all affected by the fluctuating EOF value in the electrokinetic injection mode. That is, although the analyte net migration velocity in the separation channel (CD) may remain unchanged during electrokinetic injection, the fluctuating injection speed in the sample channel (AC) can induce a changing velocity in the collection channel (BC) based on the principle of conservation of effective volumetric flow rate.
In addition to the present teachings enabling faster processing and reducing the effects of EOF instability, methods and systems in accordance with the present avoid buffer depletion that commonly occurs in known 2D-EFD devices during prolonged sample injection due to electrolysis of the sample buffer at the sample inlet from the electrode at the sample inlet. That is, whereas an electrode is directly placed into the sample vial in electrokinetic injection, methods and systems in accordance with various aspects of the present teachings do not use an electrode at the sample inlet and instead utilize pressure to drive the sample fluid (and the analyted contained therein) to the outlet junction.
Therefore, the systems and methods in accordance with the present teachings can be superior to electrokinetic injection in the continuous chemical purification process, which can provide faster sample processing, be more resistant to the fluctuating EOF, and avoid buffer depletion that is common in known 2D-EFD device.
It will be appreciated that various of the above-disclosed and other features and functions, or alternatives thereof, may be desirably combined into many other different systems or applications. It will also be appreciated that various presently unforeseen or unanticipated alternatives, modifications, variations or improvements therein may be subsequently made by those skilled in the art which alternatives, variations and improvements are also intended to be encompassed by the following claims.
This application claims priority to U.S. provisional application No. 62/030,989, filed on Jul. 30, 2014, which is incorporated herein by reference in its entirety.
Filing Document | Filing Date | Country | Kind |
---|---|---|---|
PCT/IB2015/055740 | 7/29/2015 | WO | 00 |
Number | Date | Country | |
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62030989 | Jul 2014 | US |