JP6526758B2 - Microfluidic chip - Google Patents

Description

  The present invention relates to microfluidic chip designs used to separate particles or cellular material into various components and fragments using laminar flow.

  The present invention relates to microfluidic chip designs used to separate particles or cellular material into various components and fragments using laminar flow.

[Description of related fields]
When separating various particles or cellular material, eg separating sperm from non-viable or non-motile sperm into viable motile sperm or by sex, the process is time consuming and imposes severe volume constraints. Often work. As such, current separation techniques can not, for example, timely produce the desired outcome or processing volume of cellular material.

  Thus, there is a need for separation techniques and devices that are continuous, high throughput, time-saving, and negligible or minimal damage to the various components being separated. In addition, such devices and methods can be used not only in the classification of sperm, but also in the separation of blood and other cellular material including viruses, organelles, spherical structures, colloidal suspensions and other biological material. It must have further applicability to medical and medical areas.

  The present invention relates to a microfluidic chip system, which comprises a microfluidic chip loaded into a microfluidic chip cassette mounted on a microfluidic chip holder.

  In one embodiment, the microfluidic chip comprises a plurality of layers in which a plurality of channels are arranged, the plurality of channels being a sample injection channel into which a sample fluid mixture of a plurality of components to be separated is injected; A first plurality of sheath fluid channels into which sheath fluid is introduced and intersects the sample input channel at a first intersection point, wherein the plurality of sheath fluids compress the sample fluid mixture in at least two sides, the sample fluid mixture A first plurality of sheath fluid channels and a first plurality of sheath fluid channels are relatively smaller and thinner flows defined by the plurality of sheath fluids while maintaining laminar flow in the sample input channel. With substantially the same dimensions as that of the And a second plurality of sheath fluid channels that intersect the sample input channel at a second intersection point downstream of the first intersection point in a second downward direction, the second plurality of sheath fluid channels The plurality of sheath fluids from the channel compresses the sample fluid mixture, and the plurality of components of the sample fluid mixture are compressed and oriented in a predetermined direction while further maintaining laminar flow in the sample input channel. A second plurality of sheath fluid channels and a plurality of outlet channels extending from the sample input channel, wherein the plurality of components and the plurality of sheath fluids are removed from the microfluidic chip.

  In one embodiment, the microfluidic chip includes an interrogation device that interrogates and identifies a plurality of components of the sample fluid mixture of the sample input channel in an interrogation chamber located downstream of the second intersection point .

  In one embodiment, the microfluidic chip includes a separation mechanism that separates the plurality of selected components from the plurality of components of the sample fluid mixture downstream of the interrogation chamber, which includes the sample fluid mixture in the sample input channel. By displacing the flow trajectory and pushing selected components of the displaced sample fluid mixture flow out of the interrogation chamber into one of a plurality of outlet channels extending.

  In one embodiment, the microfluidic chip comprises at least one jet chamber comprising a plurality of sheath fluids introduced into the jet chamber by at least one vent and at least one jet channel connected to the at least one jet chamber. And at least one jet channel that enters the sample input channel in the interrogation chamber.

  In one embodiment, the separation mechanism includes at least one piezoelectric actuator assembly disposed on at least one side of the sample loading channel.

  In one embodiment, the piezoelectric actuator assembly is an external laminated piezoelectric actuator assembly.

  In one embodiment, the microfluidic chip further includes a diaphragm covering each of the jet chambers, and the outer laminated piezoelectric actuator assembly drives a plurality of sheath fluids in the jet chamber into the sample loading channel and in the sample loading channel The diaphragm is aligned with and displaced so as to displace the trajectory of the sample fluid mixture flow into one of the plurality of outlet channels.

  In one embodiment, the external stacked piezoelectric actuator assembly is disposed on the microfluidic chip holder.

  In one embodiment, the microfluidic chip further includes an electronic circuit connected to the piezoelectric actuator assembly, wherein the electronic circuit amplifies an electronic signal generated by a resistive force from the piezoelectric actuator in contact with the diaphragm.

  In one embodiment, the electrical signal from the piezoelectric film is indicative of the amount of strain generated by the outer laminated piezoelectric actuator assembly.

  In one embodiment, the indicator of contact is automatically turned on when a contact is formed between the piezoelectric actuator and the diaphragm.

  In one embodiment, when contact is detected, the electronic signal exceeds a set threshold and the piezoelectric actuator assembly compresses the jet chamber to cause multiple sheath fluids to be ejected from the jet chamber into the sample fluid channel.

  In one embodiment, the indicator of touch comprises light, sound, touch or any combination thereof.

  In one embodiment, the piezoelectric actuator assembly includes a flexible diaphragm covering the jet chamber, and a piezoelectric material adhered to the top surface of the diaphragm by an adhesion mechanism.

  In one embodiment, when voltage is applied to the plurality of electrodes of the piezoelectric actuator assembly, the flexible diaphragm flexes into the jet chamber to deflect the plurality of selected components into one of the plurality of outlet channels, Of the sheath fluid from the jet chamber into the sample input channel. In one embodiment, the jet channel is tapered when it connects to the sample input channel.

  In one embodiment, the microfluidic chip further comprises a plurality of outlets disposed at a plurality of ends of the plurality of outflow channels.

  In one embodiment, the plurality of outlet channels are larger in size than the sample input channels.

  In one embodiment, the microfluidic chip further includes a plurality of notches disposed at the lower end of the microfluidic chip to separate the plurality of outlets.

  In one embodiment, the sample injection channel and the plurality of sheath channels are disposed in one or more planes of the microfluidic chip.

  In one embodiment, the sample loading channel and the plurality of sheath channels are disposed in or between one or more structural layers of the microfluidic chip.

  In one embodiment, at least one of the plurality of sheath channels is disposed in a plane different from the plane in which the sample injection channel is disposed.

  In one embodiment, at least one of the plurality of sheath channels is disposed in a structural layer different from the structural layer in which the sample loading channel is disposed.

  In one embodiment, the sample injection channel tapers at an entry point to a first intersection point with the plurality of sheath channels. In one embodiment, the sample input channel is tapered towards the interrogation chamber.

  In one embodiment, the plurality of sheath fluid channels taper at a plurality of entry points into the sample input channel at at least one of the first intersection point or the second intersection point.

  In one embodiment, the interrogation chamber includes an opening that is cut through the plurality of structural layers of the microfluidic chip, and the upper window has a first cover at the opening of at least one of the plurality of structural layers. The lower window is configured to receive, and the lower window is configured to receive the second cover at the opening of at least one of the plurality of structural layers.

  In one embodiment, the interrogation chamber includes an opening that is cut through the plurality of planes of the microfluidic chip, and the upper window is formed at a first opening of at least one plane of the plurality of planes of the microfluidic chip The lower window is configured to receive the second cover at an opening of at least one of the plurality of planes of the microfluidic chip.

  In one embodiment, the interrogation apparatus includes a light source configured to emit a beam through the first cover to illuminate and excite multiple components of the sample fluid mixture, the light induced and emitted by the beam Passes through the second cover and is received by the objective lens.

  In one embodiment, the interrogation apparatus includes a light source configured to emit a beam through the plurality of structural layers of the microfluidic chip to illuminate and excite the plurality of components of the sample fluid mixture, and is directed by the beam The emitted light is received by the objective lens.

  In one embodiment, the interrogation apparatus includes a light source configured to emit a beam through the plurality of planes of the microfluidic chip to illuminate and excite the plurality of components of the sample fluid mixture, the beam being induced to emit The received light is received by the objective lens.

  In one embodiment, the emitted light received by the objective is converted into an electronic signal that triggers the piezoelectric actuator assembly.

  In one embodiment, one of the sample fluid mixture or the plurality of sheath fluids is pumped to the microfluidic chip by a pumping device. In one embodiment, the outer tube communicates multiple fluids to the microfluidic chip. In one embodiment, the plurality of components is a plurality of cells.

  In one embodiment, the plurality of cells to be separated is a viable motile sperm from non-viable or non-motile sperm, a sperm isolated by gender and other gender classification variations, a plurality of stem cells separated from population cells, plural One or more labeled cells separated from multiple unlabeled cells, including sperm cells, multiple cells including multiple sperm cells, defined by desirable or undesirable characteristics, nuclear DNA according to particular features Separation of multiple genes separated in multiple cells, multiple cells separated based on multiple surface markers, multiple cells separated based on membrane integrity or viability, separation based on possible or predicted regeneration status Cells isolated, cells isolated based on frozen viability, cells isolated from contaminants or debris, and Plural healthy cells separated from damaged cells, plural white blood cells of plasma mixture and plural red blood cells separated from platelets, or plural corresponding fragments from any other cellular components And at least one of any cells of

  In one embodiment, the plurality of separated components are moved to one of the plurality of outlet channels, and the plurality of unselected components flow out through the other of the plurality of outlet channels.

  In one embodiment, the microfluidic chip further comprises a computer that controls the pumping of the sample fluid mixture or the plurality of sheath fluids to one microfluidic chip.

  In one embodiment, the microfluidic chip further includes a computer that displays the plurality of components in a field of view acquired by a CCD camera disposed over the aperture of the microfluidic chip.

  In one embodiment, the microfluidic chip system is a microfluidic chip loaded in a microfluidic chip cassette mounted on a microfluidic chip holder, the sample input unit for introducing a sample fluid into the microfluidic chip, A microfluidic chip having a plurality of sheath inputs for introducing a sheath fluid into the microfluidic chip, and a sample fluid is pumped from the reservoir to the sample input of the microfluidic chip, and a plurality of sheath fluids are applied to the plurality of microfluidic chips And a delivery mechanism for delivering pressure to the sheath insertion part of

  In one embodiment, a method of directing and separating components of a fluid mixture comprises: injecting a sample fluid mixture comprising the components into a sample loading channel of the microfluidic chip; Injecting a plurality of sheath fluids into the sheath fluid channel of the plurality of sheath fluids from the plurality of first sheath fluid channels, the plurality of first sheath fluid channels and a first intersection of the plurality of sample introduction channels Wherein the plurality of sheath fluids from the plurality of first sheath fluid channels merge with the sample fluid mixture in the sample loading channel to cause the plurality of components of the sample fluid mixture to be concentrated around the center of the sample loading channel Compressing the sample fluid mixture in one direction of the input channel; Injecting a plurality of sheath fluids into a number of second sheath fluid channels, the plurality of sheath fluids from the plurality of second sheath channels being downstream of the first intersection point; The plurality of sheath fluids from the plurality of second sheath fluid channels merge with the plurality of sample fluid channels, and the plurality of sheath fluids from the plurality of second sheath fluid channels merge with the sample fluid mixture of the sample loading channel The sample fluid mixture is further compressed in the second direction at the second intersection so that the plurality of components converge and align in the center of the sample input channel, both by width and depth, as it flows through. The sheath fluid acts on the plurality of components to compress and direct the plurality of components in a selected direction as the plurality of components flow through the sample fluid channel That includes a stage, a.

  Thus, some features of the present invention have been outlined in order to provide a better understanding of their detailed description, which is set forth below, and a better understanding of the present contribution to the art. Of course, the additional features according to the invention described below form the subject matter of the claims appended hereto.

  In this regard, before describing in detail at least one embodiment in accordance with the present invention, the present invention will be described in detail in the following description or its application to the details of construction and arrangement of components shown in the drawings. It should be understood that it is not limited. The methods and apparatus according to the present invention may take other embodiments and may be implemented and carried out in various ways. Also, it is to be understood that the phraseology and terminology employed herein, as well as the abstract included below, are for the purpose of description and should not be regarded as limiting.

  As such, one of ordinary skill in the art can readily utilize the concepts underlying the present disclosure as a basis for designing other structures, methods, and systems for carrying out the several purposes of the present invention. It will be understood that Therefore, it is important that the claims be regarded as including equivalent arrangements of the various methods and devices of the present invention without departing from the spirit and scope thereof.

BRIEF DESCRIPTION OF THE DRAWINGS The objects, features and advantages of the present invention will be more readily understood as the following description of the disclosure is considered in conjunction with the accompanying drawings.
FIG. 7 shows an exploded perspective view of an exemplary embodiment of a microfluidic chip according to an embodiment according to the present invention. FIG. 2A shows a top view of the assembled microfluidic chip of FIG. 1 according to a variant embodiment according to the invention. FIG. 2A shows a top view of the assembled microfluidic chip of FIG. 1 according to a variant embodiment according to the invention. FIG. 2A shows a top view of the assembled microfluidic chip of FIG. 1 according to a variant embodiment according to the invention. FIG. 3 shows a cross-sectional view of the interrogation chamber of the microfluidic chip of FIGS. 1 and 2A-2C according to an embodiment according to the present invention. Exemplary interrogation with light sources of multiple components flowing through the fluid mixture through the microfluidic chip of FIGS. 1 and 2A-2C, and two (mirror images), according to one embodiment according to the present invention FIG. 7 shows an internal cross-sectional view of one exemplary operation of a piezoelectric actuator assembly. FIGS. 2A-2C show internal perspective and perspective views of the multiple components flowing through the microfluidic chip of FIGS. 1 and 2A-2C, and an exemplary operation of a two-stage centralized, according to one embodiment according to the present invention. FIG. 1C shows a perspective view and a perspective view of a plurality of channels and interrogation chambers disposed in the microfluidic chip of FIGS. 1-2C, according to one embodiment according to the present invention. FIG. 5 shows a schematic view of a front view of a main body of a microfluidic chip holder according to an embodiment of the present invention. FIG. 7 shows a schematic view of a side view of a piezoelectric actuator assembly of the microfluidic chip holder of FIG. 6 according to an embodiment according to the present invention. FIG. 1 shows a schematic view of a front view of a microfluidic chip holder according to an embodiment of the present invention. FIG. 8 illustrates a pumping mechanism for pumping sample fluid and sheath or buffer fluid to a microfluidic chip, according to one embodiment according to the present invention.

  DETAILED DESCRIPTION Before referring to the figures that illustrate the example embodiments in detail, it is understood that the present disclosure is not limited to the details or methodology set forth in the description or illustrated in the figures. I want to be It is also to be understood that the terms are for the purpose of illustration only and should not be considered limiting. The same or similar reference numerals are intended to be used in all the drawings to refer to the same or similar parts.

  The present disclosure relates to microfluidic chip design, which is used to separate cellular material such as particles or sperm and other particles or cells into various components and fragments using laminar flow.

  Various embodiments of the present invention separate sperm from separated non-viable or non-motile spermatozoa, eg, by sex and other gender classification variations, to separate multiple components of the mixture. Separating a plurality of stem cells from a population of cells, clarifying desired / undesired properties, and separating one or more labeled cells from a plurality of unlabeled cells, a plurality of nuclear DNA according to a particular feature. Separating genes, separating multiple cells based on multiple surface markers, membrane integrity (viability), possible or predicted regeneration status (fertility), frozen viability etc. Separating multiple cells, separating multiple cells from multiple contaminants or debris, (as in bone marrow extraction) multiple damaged cells (Separating from cancerous cells), separating multiple red blood cells from multiple white blood cells and platelets of plasma mixture, multiple arbitrary cells from multiple arbitrary other cellular components and multiple corresponding fragments To provide separation.

  Furthermore, the subject matter of the present disclosure is suitable for other medical applications as well. For example, the various laminar flows described below are available as part of a renal dialysis treatment where whole blood is removed from the waste and returned to the patient. Furthermore, various embodiments of the present disclosure include cells, viruses, bacteria, organelles or subparts, spherical structures, colloidal suspensions, lipids and lipid spheres, gels, immiscible particles, blastomeres, cell aggregates, microorganisms And may have further applicability to other biological or medical areas, such as the separation of other biological materials. For example, separation of multiple components according to the present disclosure may include cell "washing" where contaminants (such as bacteria) are removed from the cell suspension and may be particularly useful in medical and food industry applications . Significantly, prior art flow-based techniques have not recognized any application to the separation of non-motor cell components as in the present invention.

  The subject matter of the present disclosure is also applicable to transferring species from one solution to another, wherein separation by filtration or centrifugation is impractical or undesirable. In addition to the applications described above, additional applications, for example, separate colloids of a given size from colloids of other sizes (for research or commercial applications) Washing the plurality of particles such as cells, egg cells etc. (effectively displacing the medium in which they are contained, removing contaminants) or of nanotubes from a solution of salt and surfactant Washing the plurality of such particles with different salt concentrations or without surfactant.

  The action of the separation of the multiple species may be a number of physical properties of the object or component, including self-moving, self-diffusing, free falling velocity, or operating under external forces such as actuators, electromagnetic fields or holographic optical traps. May vary depending on The properties on which the classification is based are, for example, cell motility, cell viability, size of the object, mass of the object, concentration of the object, the object attracts or repels each other or other objects in the flow Including the tendency, the charge of the object, the surface chemistry of the object, and the tendency of certain other objects (ie, molecules) to adhere to the object.

  Various embodiments of the plurality of microfluidic chips utilize one or more flow channels, as described below, which have a plurality of substantially laminar flows and one or more components are Interrogated for identification, enabling it to be separated into multiple streams flowing out to one or more outlets. Furthermore, the various components of the mixture are separated on-chip, for example by using flow mechanisms or additional separation mechanisms such as optical tweezers or holographic optical traps, or by magnetism (ie utilization of magnetic beads) It may be done. Various embodiments of the present invention thereby allow multiple components to be defined in a continuous closed system without the possibility of damage and contamination as in the prior art methods, in particular in sperm separation. To achieve continuous separation. The continuous processing of the present invention further realizes significant time savings when separating multiple components.

  The following description is for separating sperm from non-viable or non-motile sperm into viable motile sperm, or separating sperm by gender and other gender classification variations, or clarifying desirable / undesired characteristics etc. Or focusing on separating multiple labeled cells from multiple unlabeled cells, the devices, methods and systems of the present invention can be interrogated by fluorescence techniques in a fluid stream, or different to different effluents It may be extended to other types of particulate matter, live matter or cellular matter operable between fluid streams.

  Although the present subject matter will be described in detail with respect to the microfluidic chip 100 shown in FIGS. 1-5 and the microfluidic chip holder 200 shown in FIGS. 6-9, the description may be made in various ways as described herein. It should be understood that the present invention applies equally to the other embodiments or any of these variations.

[Microfluidic chip assembly]
FIG. 1 is an exemplary embodiment of a microfluidic chip 100. The microfluidic chip 100 is manufactured from an appropriate thermoplastic (eg, low auto-fluorescent polymer, etc.) through embossing or injection molding processes, as is well known to those skilled in the art, and is of a suitable size.

  The microfluidic chip 100 includes a plurality of structural layers, in which a plurality of microchannels functioning as a sample input channel, a sheath or buffer fluid channel, an outflow channel, etc. are arranged. The plurality of microchannels are of a size suitable to accommodate the laminar particle flow, and in any of the plurality of layers of the chip 100, arranged with an appropriate length as long as the object of the present invention is realized. It is also good. The desired flow rate through the microfluidic chip 100 can be achieved by pumping mechanisms, providing narrowing at various locations of the plurality of microchannels, and / or providing obstacles or partitions within the plurality of microchannels. , May be controlled by a predetermined introduction flow rate into the chip 100 while maintaining the appropriate dimensions of the microchannels in the chip 100.

  A plurality of inlets are provided on the microfluidic chip 100, which provide access to the plurality of microchannels / channels. In one embodiment, as shown in FIG. 1 and FIGS. 2A-2C, the sample input 106 may store a sample of the plurality of components 160 of the sample fluid mixture 120 (see FIGS. 4-5). Is used to introduce the sample into the sample input channel 164A of the microfluidic chip 100. The microfluidic chip 100 further includes at least one sheath / buffer input (in one embodiment, sheath / buffer inputs 107, 108) for introducing a sheath or buffer fluid. In one embodiment, there are two sheath / buffer inputs in the microfluidic chip 100, including a sheath / buffer input 107 and a sheath / buffer input 108, both of which are located near the sample input 106, Both introduce a sheath or buffer fluid into the microfluidic chip 100. Sheath or buffer fluids are well known in the microfluidic art and, in one embodiment, well known in the art to maintain the viability of multiple components 160 (ie, multiple sperm cells) of the fluid mixture. It may contain the nutrients of The position of the sheath / buffer loaders 107, 108 may be different and they may access multiple microchannels in the same or different structural layers of the chip 100.

  In one embodiment, the fill holes or vents 121, 122 may be used to introduce a sheath or buffer fluid into the jet chamber 130, 131 (described below) if not sealed.

  In one embodiment, a plurality of outlet channels extending from the main channel 164 (see FIG. 2A) are provided to remove fluid comprising the separated component 160 and / or sheath or buffer fluid that has flowed through the microfluidic chip 100. Be In one embodiment, as shown in FIGS. 1 and 2A-2C, there are three outflow channels 140-142 including a left outflow channel 140, a central outflow channel 141 and a right outflow channel 142. The left outflow channel 140 terminates at the first outflow 111, the central outflow channel 141 terminates at the second outflow 112, and the right outflow channel 142 terminates at the third outflow 113. However, the number of outlets may be small or large, depending on the number of components 160 to be separated from the fluid mixture 120.

  In one embodiment, instead of straight ends, where necessary, a plurality of notches or recesses 146 separate a plurality of outlets (i.e. outlets 111-113), an outer pipe or the like. Located at the lower end of the microfluidic chip 100 for attachment. The first outlet 111, the second outlet 112 and the third outlet 113 are reached via the outlet channels 140-142 originating from the interrogation chamber 129 (see FIG. 2A-4).

  In one embodiment, the microfluidic chip 100 has a plurality of structural layers in which a plurality of microchannels are arranged. Multiple channels may be disposed in one or more layers, or between multiple layers. In one embodiment, as shown in FIG. 1, by way of example, four structural plastic layers 101-104 are shown as comprising a microfluidic chip 100. However, one skilled in the art will realize that fewer or additional layers may be used and that multiple channels may be located in any of multiple layers as long as the objects of the present invention are achieved. Will.

  A gasket or o-ring of any desired shape may be provided to maintain a tight seal between the microfluidic chip 100 and the microfluidic chip holder 200 (see FIG. 6). In the case of a gasket, this may be a single sheet or multiple components of any configuration or material (i.e., rubber, silicone, etc.), as desired. In one embodiment, as shown in FIG. 1, a first gasket 105 is disposed at one end of the microfluidic chip 100, bonded to the layer 104, or bonded thereto (using an epoxy) Ru. A plurality of holes 144 are provided in the first gasket 105 and aligned with the sample input 106, the sheath / buffer input 107, the sheath / buffer input 108 and the vents 121, 122 to provide access thereto. Configured to

  In one embodiment, the second gasket 143 is disposed at the other end of the microfluidic chip 100 opposite the first gasket 105 and combines with the upper structural layer 104 or (with epoxy) Glued to. The second gasket 143 is configured to help seal, stabilize or balance the microfluidic chip 100 in the microfluidic chip holder 200 (see FIG. 6).

  In one embodiment, the plurality of holes and the plurality of posts 145 are disposed at various convenient locations on the microfluidic chip 100 to fix and align the plurality of layers (ie, layers 101-104) during chip processing. Ru.

  In one embodiment, a sample fluid mixture 120 comprising a plurality of components 160 is introduced into the sample input 106, and the fluid mixture 120 passes through the main channel 164 to the interrogation chamber 129 (see FIGS. 2A, 4 and 5). Flow toward The sheath or buffer fluid 163 is introduced to the sheath / buffer input 107, 108 and flows through the channels 114, 115 and 116, 117 to the main channel 164 and out through the plurality of outflow channels 140-142, respectively. Head to the interrogation chamber 129 forward.

  In one embodiment, if the chambers 130, 131 are not filled with a sheath or buffer fluid 163 during manufacture, the sheath or buffer fluid 163 ventilate to fill the chambers 130, 131 after the fabrication of the microfluidic chip 110. It may be introduced into the jet chamber 130, 131 through the ports 121, 122. As mentioned above, the sheath or buffer fluid 163 used is well known to those skilled in the art of microfluidics.

  In one embodiment, the fluid mixture 120 from the main channel 164 merges with the sheath or buffer fluid 163 from the channels 114, 115 at the intersection point 161 of the same plane of the microfluidic chip 100. In one embodiment, the buffer fluid 163 from the channels 116, 117 merges with the mixed fluid mixture 120 and the sheath or buffer fluid 163 from the first intersection point 161 downstream of the second intersection point 162. In one embodiment, the channels 114, 115 are substantially the same size as the channels 116, 117 as long as the desired flow rate is achieved to achieve the objects of the present invention.

  In one embodiment, channels 114-117, 123, 124, 140-142, 125a, 125b, 126a, 126b, 127, 128 may have substantially the same dimensions, however, as one of ordinary skill in the art For example, the size of any or all of the plurality of channels of the microfluidic chip 100 may be different (ie, between 50 and 500 microns) as long as the desired flow rate is achieved to achieve the objects of the present invention. It will be appreciated that there may be.

  In one embodiment, the channels 114-117, 123, 124, 140-142, 125a, 125b, 126a, 126b, 127, 128 of the microfluidic chip 100 may not only differ in size, but also a plurality of channels. It may have a tapered shape at the entry point to the plurality of other channels of the chip 100 to control the flow of fluid therethrough. For example, the main channel 164 may be tapered to control the flow of the sample 120 to the intersection point 161 at the entry point to the intersection point 161 (see FIG. 5B) and to increase its speed. Sample fluid in a first direction (i.e., horizontal) on at least two sides if not all sides (depending on where the tapered channel 164 joins the channel 164A) the sheath or buffer fluid 163 from It may be possible to compress the mixture 120. That is, the sample fluid mixture 120 will be a relatively smaller, thinner stream defined or surrounded by the sheath or buffer fluid 163 while maintaining the laminar flow of the channel 164A. However, one skilled in the art will be aware that the main channel 164 entering the intersection point 161 may have any physical configuration, such as a rectangular or circular channel, as long as the object of the present invention is obtained.

  In an exemplary embodiment, at least one of the channels 116, 117 is disposed in the microfluidic chip 100 in a structural layer different from the layer in which the channel 164 is disposed. For example, if the sheath or buffer fluid 163 joins the fluid mixture 120 at the intersection point 162, the channel 116 is in a different plane from the other channels 164 and 114, 115 (in the layer 102), It may be disposed in layer 103 and channels 117 may be disposed in layer 101 (see FIG. 1). In one embodiment, the main channel 164 is disposed between the layers 102, 103 (see FIG. 3), however, one skilled in the art would consider the channels 114-117, 164, 123, 124, 140-142, It will be appreciated that 125a, 125b, 126a, 126b, 127, 128 etc may be arranged in any layer or between any two layers. Further, the channels 114-117, 164, 123, 124, 140-142, 125a, 125b, 126a, 126b, 127, 128 etc are described in the exemplary embodiments as shown in the figures. However, one of ordinary skill in the art would be aware that the particular configuration or layout of the multiple channels of chip 100 may be any desired configuration as long as the described features of the invention are realized. I will.

  In one embodiment, the sheath or buffer fluid of the channels 116, 117 merges into the fluid mixture at substantially vertical positions above and below the intersection point 162 through the plurality of holes cut in the layers 101-103. . The sheath or buffer fluid from the channels 116, 117 allows laminar flow of the channel 164B so that the components 160 of the fluid mixture 120 are compressed or crushed and directed in a selected or desired direction (see below) While the flow of fluid mixture 120 is compressed perpendicular to channel 164B.

  In one embodiment, as shown in FIGS. 1 and 2A-2C, the channels 114, 115 and 116, 117 are partially coaxial with one another with respect to the central point defined by the sample input 106. As illustrated. That is, in one embodiment, the channels 114, 115 and 116, 117 are arranged in a substantially parallel configuration with the channels 114, 115 and 116, 117 equidistant from the main channel 164. However, one skilled in the art will recognize that the illustrated configurations may differ as long as they achieve the desirable features of the present invention.

  Furthermore, in one embodiment, the channels 114, 115 preferably meet at the intersection point 161 of the same plane at an angle of 45 degrees or less and the channels 116, 117 parallel to the sample injection channel 164A are from different layers , Join the intersection point 162 at an angle of substantially 90 degrees. However, one skilled in the art will appreciate that the illustrated configurations, angles, and structural configurations of the multiple layers and multiple channels of the microfluidic chip 100 are those that embody desirable features of the present invention. It will be appreciated that they may differ as long as

  In one embodiment, downstream of the intersection point 162, the plurality of components 160 of the fluid mixture 120 flow through the channel 164B to the interrogation chamber 129, where the plurality of components 160 are interrogated.

  In one embodiment, a flexible diaphragm 170, 171 formed of a suitable material, such as stainless steel, brass, titanium, nickel alloys, polymers or any other suitable material having a desirable elastic response (see FIG. 1). ) Covers the jet chamber 130, 131). In one embodiment, the actuator is disposed on at least one side of the channel 164B and the interrogation chamber 129 (see FIGS. 2A and 2B) to mechanically displace the diaphragm 170, 171 and to provide a sheath or buffer fluid. 163 is jetted or pushed out of one of the jet chambers 130, 131 at the relevant side of the channel 146B, and a plurality of components 160 from the channel 164C to one of the outflow channels 140, 142 at the other side of the channel 164B. Push out. In other words, the actuator ejects the sheath or buffer fluid 163 from the jet chamber 130 into the channel 164 C and separates the plurality of target components 160 of the channel 164 C to separate the plurality of target components from the fluid mixture 120. Push out. This embodiment is useful when only one type of target component 160 is separated (e.g. only two outflow channels 141, 142 are needed instead of three outflow channels 140-142) Also good) (see Figure 2B).

  The actuator may be one of a piezoelectric, magnetic, electrostatic, hydraulic or pneumatic actuator. Although a disc-shaped actuator assembly (ie, 109, 110) is shown in FIGS. 1 and 2A-2C, one of ordinary skill in the art can utilize any type or shape of actuator that performs the required function. It will be understood that it is.

  In other embodiments, the actuators are located on either side of the channel 164B (as shown in FIG. 2A), but in other embodiments more than one (of relatively small size) An actuator may be disposed on one or more sides of channel 164B and connected to channel 164B via a jet channel (see FIG. 2C).

  The following description of the function of the actuator is made with reference to FIG. 2A, but those skilled in the art will appreciate that any of the features placed on the chip 100 can be placed as long as they realize the multiple features of the invention It will be appreciated that types of actuators are applicable.

  In one embodiment, two outer laminated piezoelectric actuator assemblies 209, 210 are provided to activate the diaphragms 170, 171 and eject the sheath or buffer fluid 163 from the chambers 130, 131 into the channel 164B (see FIGS. 6 and 7) ), Align with the diaphragms 170, 171 and activate them. The external stacked piezoelectric actuator assemblies 209, 210 are disposed on the microfluidic chip holder 200. Each of the stacked piezoelectric actuator assemblies 209, 210 includes a piezoelectric actuator 219, 220, respectively, which have high resonant frequencies, each of which pushes the sheath or buffer fluid 163 from the chamber 130, 131 into the channel 164C. , Located at the center position of the diaphragms 170 and 171 and in contact with them.

  The microfluidic chip holder 200 may be of any type known to those skilled in the art, such that the piezoelectric actuators 219, 220 can maintain constant contact with the diaphragms 170, 171 of the microfluidic chip 100. It is configured to accurately position 219, 220. For example, in one embodiment, this is accomplished by each of the piezoelectric actuator assemblies 209, 210, which can be a plurality of lockable adjustment screws 201 that move the piezoelectric actuators 219, 220 into position relative to the diaphragms 170, 171, respectively. And a plurality of thumbscrews 202 having threaded bodies that act to move the plurality of screws 202 relative to the diaphragm 170, 171 for stabilization (or bonding using a suitable epoxy) Be done. The spacer 203 attached to the piezoelectric actuator 219, 220 enables a viable contact between it and the diaphragm 170, 171 of the microfluidic chip 100. The plurality of adjustment screws 201 allow the user to make adjustments to the position of the piezoelectric actuators 209, 210 both to provide coarse or fine adjustments to the microfluidic chip 100. The plurality of thumb screws 202 may be tightened to secure the piezoelectric assembly 209, 210 relative to the main chip body 100, and may be loosened to remove the piezoelectric actuator assembly 209, 210 from the main chip body 100. Good.

  In one embodiment, at least one piezoelectric actuator (209 or 210) is mounted on a plate (not shown), which is translatable in a direction orthogonal to the diaphragm (170 or 171) of the microfluidic chip 100 is there. The adjustment screw 201 is mounted on the holder 200, and can be expanded and contracted by rotating the screw 201. The tip of the adjustment screw 201 is in contact with the plate. As the screw 201 is extended, the plates along the piezoelectric actuators 209, 210 are translated towards the diaphragms 170, 171 in translation so that a viable contact is created between the piezoelectric actuators 209, 210 and the diaphragms 170, 171. It is pushed out. In this way, the positioning of the piezoelectric actuators 209, 210 is adjusted only through the translation of the piezoelectric actuators 209, 210, but in the previous embodiment where the piezoelectric actuators 209, 210 are mounted directly on the adjusting screw 201, The positioning of the actuators 209, 210 is a combination of translation and rotation of the piezoelectric actuators 209, 210, during which damage to the delicate piezoelectric actuators 209, 210 can occur.

  In another embodiment, the electronic circuit is connected to the laminated piezoelectric actuator assembly 209, 210 prior to actuation thereof. When each of the laminated piezoelectric actuators 219, 220 contacts each of the diaphragms 170, 171, the resistance from the diaphragms 170, 171 causes distortion in the laminated piezoelectric actuators 219, 220 that generate electronic signals. That is, the electronic circuit can amplify the electronic signal to a predetermined value that triggers the LED (light emitting diode). When the laminated piezoelectric actuator 219, 220 contacts the diaphragm 170, 171, the LED is automatically turned on to indicate that contact has occurred between the laminated piezoelectric actuator 219, 220 and the diaphragm 170, 171. This contact sensing allows sufficient force for the actuators 219, 220 to compress the chambers 130, 131 to cause the fluid 163 to be expelled into the channel 164B.

  It will be apparent to one skilled in the art that LEDs are an example of a touch indicator. For example, once contact occurs and the electronic signal exceeds the set threshold, feedback to the user is generated, which may be of the following types: light (ie LED), sound (ie buzzer), It may be tactile (i.e. vibrator) or any combination of these. That is, the user can stop adjusting the touch and maintain the touch. Of course, in one embodiment, the process described above may be automated.

  In an alternative embodiment, instead of at least one external laminated piezoelectric actuator assembly, each of the diaphragms 170, 171 is displaced (flexed) to drive multiple fluids of each of the jet chambers 130, 131 into the channel 164C, respectively. In order to form at least one piezoelectric actuator assembly 109, 110 (see FIGS. 2A and 4), a thin film of piezoelectric material (well known to those skilled in the art) is placed directly on top of at least one diaphragm 170, 171. Be done. The piezoelectric material is permanently adhered to the aforementioned flexible diaphragms 170, 171 by an adhesion mechanism. That is, in the present embodiment, when a voltage is applied to the plurality of electrodes 109, 110 of the piezoelectric actuator assembly, the entire diaphragm 170, 171 flexes into the chambers 130, 131 and fluid 163 therein into the channel 164C. The pushing diverts the plurality of target or selected components 160 towards the lateral outflow channels 140, 142.

  As noted above, in one embodiment, only one piezoelectric actuator assembly, as shown in FIG. 2B, for either the external laminated piezoelectric actuator assembly 209, 210 or the piezoelectric actuator assembly 109, 110. May be required to eject a sheath or buffer fluid 163 from the jet chamber 130 into the channel 164C and push the plurality of target components 160 of the channel 164C into the outflow channel 142 in order to separate the target components of the fluid mixture 120 from .

  In one embodiment, the piezoelectric actuator assembly 109, 110 may be used to seal each of the jet chambers 130, 131, for example, in the layer 103, although one skilled in the art would consider the chambers 130, 131 It will be appreciated that any structural layer may be such that the microfluidic chip 100 is not permeable to fluid leaks after being filled with a sheath or buffer fluid 163.

  That is, in contrast to the large displacements and strong forces exerted by the external laminated piezoelectric actuator assembly 209, 210 that allow operation at very high flow rates, the piezoelectric actuator assembly 109, 110 is relatively relative to the diaphragm 170, 171. Given the small deflection displacement and the small stress that is applied to it, the low flow velocity requirement is met. However, one of ordinary skill in the art would be aware that the actuator assemblies 109, 110, 209, 210 can be independently selected for use with the microfluidic chip 100 based on different operating speed and flow rate requirements. .

  In one embodiment, the piezoelectric thin films disposed on the diaphragms 170, 171 are produced when the outer laminated piezoelectric actuator assemblies 209, 210 are triggered by the electronic signal to displace each of the diaphragms 170, 171. Act as a strain sensor to determine the amount of strain or displacement. The diameter and thickness of the piezoelectric thin film differ depending on the cross section of the external laminated piezoelectric actuator 219, 220 and the force generated on the diaphragm 170, 171. The piezoelectric thin film and diaphragms 170, 171 may differ from those described above in alternative embodiments.

  The jet chambers 130, 131 will now be described. In one embodiment, vents 121, 122 are filled with a sheath or buffer fluid 163 after manufacture, and chambers 130, 131 are vented therein when venting air through vents 121, 122. It is provided to remove air from each of the jet chambers 130, 131 (see FIG. 2A) before being sealed with the sheath or buffer fluid 163. Alternatively, in another embodiment, if the vents 121, 122 remain open, the sheath or buffer fluid 163 is introduced into the chambers 130, 131 through the ports 121, 122, unless it is done during manufacture. It is also good. The sheath or buffer or other fluid 163 disposed in the jet chamber 130, 131 may be the same as or different from the sheath or buffer fluid 163 introduced through the channels 114, 115, 116 or 117.

  In one embodiment, when a sheath or buffer fluid 163 is used to fill the jet chambers 130, 131, they are injected through the inlets 121, 122 and flow through the channels 123, 124, respectively, into the channels 125a and 125b. It may enter jet chamber 130 via and through jet channels 131 via channels 126a and 126b.

  In one embodiment, jet channel 127 leaves jet chamber 130, jet channel 128 leaves jet chamber 131, and both jet channels 127, 128 enter interrogation chamber 129 (see FIG. 2A). The jet channels 127, 128 may be arranged in any layer of the tip 100 and enter the channels 164C at any angle in the same plane.

  In one embodiment, the jet channels 127, 128 may be tapered if they connect with the main channel 164C to form a strong instantaneous jet. However, one skilled in the art will recognize that jet channels 127, 128 may have particular angles or different configurations as long as they implement the described features of the present invention.

  In one embodiment, jet channels 127, 128 act to displace or deflect diaphragms 170, 171, respectively, to eject or push sheath or buffer fluid 163 into channel 164C. However, when the diaphragms 170, 171 return to a neutral (non-flexing) position, the jet channels 127, 128 resulting from the jet chambers 130, 131 maintain the net volume of fluid from the jet chambers 130, 131 to the channel 164C. Acts as a diffuser to ensure that the chambers 130, 131 are easily refilled with a sheath or buffer fluid 163.

  In one embodiment, the outflow channels 140-142 originate from the channel 164C in the interrogation chamber 129 to the outflows 111-113. As mentioned above, in one embodiment, more than one on-chip piezoelectric actuator assembly 109, 110 (or of any size or position) or external laminated piezoelectric actuator assembly 209, 210 jets a sheath or buffer fluid 163. It may be used to connect with each of the jet channels 127, 128 to provide additional power to cause the channels 164C to eject from the chambers 130, 131. In one embodiment, the distance from each of the inlets to the channel 164C of the jet channels 127, 128 to each of the outlet channels 140-142 is such that the plurality of target components 160 are mixed with the plurality of undesirable components 160 (described further below). The distance between the plurality of components 160 must be shorter to prevent this. In one embodiment, the cross-sections and lengths of the outflow channels 140-142 are predetermined volume fractions (ie, 2: 1: 2 or 1: 2: 1) to obtain the desired hydraulic resistance of the outflow channels 140-142. Etc) must be maintained.

  In one embodiment, the interrogation device is located downstream of the point where channels 116, 117 enter channel 164B. In one embodiment, channel 164 B tapers to interrogation chamber 129 to accelerate the flow of fluid mixture through interrogation chamber 129. However, one skilled in the art will be aware that the channels 164B need not be tapered, but may be of any size and size, as long as the present invention is performed in accordance with the desired requirements.

  The interrogation device is used to interrogate and identify the plurality of components 160 of the fluid mixture in channel 164B passing through the interrogation chamber 129. It should be noted that the channels 164B may be disposed in a single layer (ie, layer 102) or may be disposed between multiple layers (ie, layers 102, 103). In one embodiment, the interrogation chamber 129 includes an opening or window 149 (see FIG. 3), which is cut into the microfluidic chip 100 at least at the top layer (ie, layer 104 or otherwise) and the other The openings or windows 152 are cut into the chip 110 at least at the bottom layer (ie, layer 101 or otherwise).

  In one embodiment, the openings 150 are cut into the microfluidic chip through layers 101-104. In one embodiment, the upper window 149 is configured to receive the first cover 133 and the lower window 152 is configured to receive the second cover 132. However, the windows 149, 152 may be located on any suitable layer, and need not be on the top / bottom layer. The covers 133, 132 may be made of any material having the desired transmission requirements, such as plastic, glass or even lenses. Although the relative diameters of the windows 149, 152 and the openings 150 are shown in FIG. 3, they may be different according to manufacturing considerations.

  In one embodiment, the first and second covers 133, 132 described above are configured to surround the interrogation chamber 129. The windows 149, 152 and the covers 133, 132 (see FIG. 3) allow a plurality of components 160 to flow through the fluid mixture 120 of the channel 164B (see FIG. 5A) through the interrogation chamber 129 through the opening 150 It can be acted upon by a suitable light source 147 configured to emit a high intensity beam 148 having any wavelength that matches the excitable component in the mixture 120. Although laser 147 is shown, any suitable other light source that emits a beam that excites a component, such as a light emitting diode (LED), an arc lamp, etc., is available.

  In one embodiment, the high-intensity laser beam 148 from a suitable laser 147 at a preselected wavelength, such as a 355 nm continuous wave (CW) (or quasi-CW) laser 147, comprises a plurality of components 160 of the fluid mixture (ie, It is required to excite multiple sperm cells). In one embodiment, the laser 147 (see FIG. 3) passes through the window 149 of the layer 104, through the top cover 133 of the chip 100, through the opening 150, and through the cover 132 of the layer 101 of the chip 100 and the window 152 A laser beam 148 is emitted to illuminate the plurality of components 160 flowing through the channel 164B in the interrogation region 129 of the chip 100.

  In one embodiment, the light beam 148 may be provided to the plurality of components 160 by an optical fiber incorporated into the aperture 150 of the microfluidic chip 100.

  The high-intensity beam 148 interacts with a plurality of components 160 (see detailed description below) and emits emitted light 151 directed by the beam 148 through the first and second covers 133, 132. Comes out of the lower window 152 so as to be received by the objective lens 153. The objective lens 153 may be disposed at any suitable position relative to the microfluidic chip 100. Because the interrogation chamber 129 is sealed by the first and second covers 133, 132, the high intensity beam 148 does not act on the microfluidic chip 100 and does not damage the layers 101-104. That is, the first and second covers 133, 132 help to avoid damage to the microfluidic chip 100 due to the high intensity beam 148 and photonic noise induced from the microfluidic chip material (ie, plastic).

  In one embodiment, the emitted light 151 received by the objective lens 153 is converted into an electronic signal by an optical sensor 154 such as a photomultiplier tube (PMT) or a photodiode. The electronic signals may be digitized by an analog to digital converter (ADC) 155 and sent to a digital signal processor (DSP) based controller 156. The DSP-based controller 156 monitors the electronic signals and then two actuator drivers (ie, 157a, 157a) to drive the associated one of the two piezoelectric actuator assemblies (109, 110 or 209, 210). One of 157b) may be triggered with a predetermined value.

In one embodiment (shown in FIG. 2A), the plurality of piezoelectric drivers and the plurality of piezoelectric actuators (158a, 158b or 219, 220) are respectively one of two piezoelectric actuator assemblies (109, 110 or 209, 210). And located on either side of the interrogation chamber 129. Trigger signals sent to multiple piezoelectric actuators (109, 110 or 219, 220) activate specific piezoelectric actuator assemblies (109, 110, 209, 210) when multiple selected components are detected To be determined by the sensor raw signal.

  In embodiments having bonded piezoelectric actuator assemblies 109, 110, the thickness of diaphragms 170, 171 may be different, depending on the voltage applied through the plurality of wires through actuator assemblies 109, 110 of tip 100. It is different. When electronic signals are transmitted directly to the plurality of actuator assemblies (ie, 109, 110) through the electronic circuitry, the diaphragms 170, 171 flex and change (increase) the pressure in the chambers 130, 131.

  Since at least one of the plurality of piezoelectric actuator assemblies (109, 110 or 209, 210) leaves the opening 150 to the interrogation area 129 after the interrogation, the plurality of fluid mixtures in the channel 164C Is used to act on the desired component 160 of Although the actuator driver 157b and the piezoelectric actuator assembly 110 are not shown in FIG. 4, the operation and configuration of the actuator driver 157b and the piezoelectric actuator assembly 110 are the same as those of the actuator driver 157a and the piezoelectric actuator assembly 109. That is, the piezoelectric actuator 157 b acts to deflect the plurality of components 160 of the flow of the channel 164 C towards the right outflow channel 142 and towards the third outflow 113. The same action is applied to the piezoelectric actuator assembly 110 which causes the sheath or buffer fluid 163 to be ejected from the jet chamber 131 via the jet channel 128 and the plurality of target or selected components 160 to be left outflow channel 140 and the third To the outlet 113 of the

  In an alternative embodiment, a piezoelectric actuator assembly 106A (i.e., a suitably sized 109, 110 piezoelectric disc similar to a piezoelectric actuator assembly; see FIG. 2C) or a suitable pumping system (see FIG. 9, for example, described below) Are used to pump the sample fluid 120 of the channel 164 towards the point of intersection 161. The sample piezoelectric actuator assembly 106A is disposed at the sample launching station 106. By pumping the sample fluid mixture 120 into the main channel 164, the means of control are arranged such that when the plurality of components 160 enter the main channel 164, a controlled relationship between them can be formed. A plurality of components 160 may be formed with respect to space.

  If the piezoelectric actuator assembly 109, 110 is not used, multiple (target) components 160 travel from the main channel 164 to the central outflow channel 141 and to the second outflow 112, and the sheath or buffer fluid 163 flows out. Proceed through channels 140, 142 to outlets 110, 112, respectively.

  In one embodiment, the outflow channels 140-142 leave the interrogation chamber 129 from the channel 164C and increase in size such that the outflow rate for concentration of the plurality of separated components 160 is increased through the associated channel.

[Operation of chip]
In one embodiment, the microfluidic chip 100 may be provided in a sterile state and may be provided with one or more solutions (ie, a sheath or buffer fluid 163), or according to a plurality of known methods. It may be cleaned from any fluid or materials either by draining the tip 100 or flowing a sheath or buffer fluid 153 or other solutions through the microfluidic chip 100. Once the microfluidic chip 100 is prepared and the jet chamber 130, 131 is filled with the sheath or buffer fluid 163, the vents 121, 122 can be sealed either at the time of manufacture or later (as described above). It is stopped. As mentioned above, in other embodiments, the vents 121, 122 may remain open so that additional sheath or buffer fluid 163 may be added to the chambers 130, 131 during operation.

  In one embodiment, as described above, the plurality of components 160 to be separated separates spermatozoa, for example by separating viable motile sperm from non-viable or non-motile spermatozoa, gender and other gender classification variations Separating the plurality of stem cells from the population of cells, defining the plurality of desired / undesired properties, and separating the one or more labeled cells from the plurality of unlabeled cells, having different desired properties. Separation of multiple sperm cells, genes of nuclear DNA according to specific features, separation of multiple cells based on multiple surface markers, membrane integrity (viability), possible or predicted regeneration status Separating multiple cells based on (fertility), freeze viability etc., separating multiple cells from multiple contaminants or debris, multiple Separating multiple healthy cells from damaged cells (i.e., cancerous cells) (as in bone marrow extraction), multiple white blood cells of plasma mix and multiple red blood cells from platelets, multiple any other cells It includes separating any plurality of cells from components into corresponding fragments, damaged cells or contaminants or debris, or any other biological material that it is desired to be separated. The plurality of components 160 may be a plurality of cells or a plurality of beads treated or coated with a plurality of linker molecules, or incorporated into a fluorescent or luminescent labeled molecule. The plurality of components 160 may have various physical or chemical properties such as size, shape, material, feel, etc.

  In one embodiment, heterogeneous populations of multiple components 160 may be measured simultaneously, wherein each component 160 is tested (eg, multiplexed measurements) for different amounts or types in similar amounts, or The plurality of components 160 may be labels (eg, fluorescence), images (size, shape, different absorption, scattering, fluorescence, emission characteristics, fluorescence or emission emitting statistics, fluorescence or emission decay period), and / or particle position etc. May be examined and clarified on the basis of

  In one embodiment, the two-stage centralized method of component classification system according to the present invention, as shown in FIG. 5A, comprises a plurality of components 160 for channel 164 B for interrogation in interrogation chamber 129. May be used to position the

  In one embodiment, the first concentration step of the present invention comprises loading a fluid sample 120 comprising a plurality of components 160, such as a plurality of sperm cells or the like, through the sample input 106, and a sheath or buffer fluid 163. , Or through a sheath or buffer loader 107,108. In one embodiment, the plurality of components 160 are pre-colored with a dye (eg, a Hoechst dye) to allow fluorescence and for image analysis to be detected. In one embodiment, a sheath or buffer fluid 163 is disposed in the jet chambers 130, 131 and the inputs 121, 122 are sealed.

  In one embodiment, as shown in FIG. 5A, multiple components 160 of the sample fluid mixture 120 flow through the main channel 164 and assume a random orientation and position (see inset A). At intersection point 161, sample mixture 120 flowing through main channel 164 is in a first direction (ie, at least the first direction if sheath or buffer fluid 163 merges with sample mixture 120 by sheath or buffer fluid 163 from channels 114, 115). Horizontally, it is compressed at least on both sides of the flow, if not on all sides depending on the position where the main channel 164 enters the intersection point 161. As a result, multiple components 160 may be concentrated around the center of channel 164 and compressed into flakes across the depth of channel 164A. An intersection point 161 reaching the channel 164A is a concentration area. That is, when sample 120 is compressed toward the center of channel 164 A by sheath or buffer fluid 163 from channels 114, 115 at intersection point 161, multiple components 160 (ie, multiple sperm cells) Move towards the middle of the width of the.

  In one embodiment, the present invention includes a second concentration step, wherein the sample mixture 120 comprising the plurality of components 160 enters the channels 116, 117 at the intersection point 162 by the sheath or buffer fluid 163. It is further compressed from the direction of (ie, vertically from top to bottom). The intersection point 162 reaching the channel 164B is a second concentration area. Although the multiple inlets from the channels 116, 117 to the intersection point 162 are shown as rectangular, one skilled in the art will understand that any other suitable configuration (i.e., a taper, a circle) may be used. Please note that it will. The sheath or buffer fluid 163 of the channels 116, 117 (which may be arranged in different layers of the microfluidic chip 100 from the channels 164A-B) may be multiple in the middle of the channel 164B by both width and depth (ie horizontal and vertical) The channels 164A-B in different planes as they flow along the channel 164B to align the components 160 of the

  That is, in one embodiment having the second concentration step of the present invention, the sample mixture 120 is recompressed by the vertical sheath or buffer fluid 163 entering the channels 116, 117 and the flow of sample 120 is shown in FIG. 5A. As shown, centered at the depth of channel 164B, the plurality of components 160 flow along the center of channel 164B in an approximately single-row fashion.

  In one embodiment, the plurality of components 160 is a plurality of sperm cells 160 and is a pancake-shaped or flat tear-shaped head, such that the plurality of sperm cells 160 undergo a second concentration step That is, the plurality of flat surfaces perpendicular to the direction of light ray 148 (see FIG. 5A) redirect itself in a predetermined direction. That is, a plurality of sperm cells 160 establish a preference for their body orientation while passing through a two-step centralized process. Specifically, multiple sperm cells 160 tend to be more stable due to their flat bodies being perpendicular to the direction of compression. Thus, by control of the sheath or buffer fluid 163, the plurality of sperm cells 160 starting in a random orientation now achieve a uniform orientation. That is, the plurality of sperm cells 160 not only form a single row at the center of the channel 164B, but they further achieve a uniform orientation due to their flat surface orthogonal to the compression direction in the second concentration stage.

  That is, although all the components 160 introduced into the sample input part 106 may be other types of cells or other materials as described above, the plurality of components 160 pass through the channel 164B in a single row format. Allowing for easier interrogation of the components 160, subject to a two-step concentration step that allows to move in a more uniform orientation (depending on the type of components 160).

  In one embodiment, further downstream of the channel 164B, the plurality of components 160 are detected through the covers 132, 133 at the opening 150 of the interrogation chamber 129 using the light source 147. The light source 147 emits a light ray 148 (which may be via an optical fiber) which is centered at the aperture 150 at the center of the channel 164C. In one embodiment, the plurality of components 160, such as the plurality of sperm cells 160, have a flat surface of the plurality of components 160 by the plurality of concentrated flows (ie, the flow of the sheath or buffer fluid 163 also acting as the sample flow 120). It is directed to point at beam 148. In addition, all components 160 are moved in a single row fashion by centralization as they pass under beam 148. As components 160 pass under light source 147 and are acted upon by beam 148, components 160 emit fluorescence that is indicative of a plurality of desirable components 160. For example, for multiple sperm cells, multiple X chromosome cells fluoresce at different intensities than multiple Y chromosome cells, or multiple cells with one property may have different sets of multiple properties. It may emit fluorescence at a different intensity or wavelength than a plurality of cells having. Further, the plurality of components 160 may be viewed for shape, size or any other plurality of clarification indicators.

  In the beam induced fluorescence embodiment, the emitted light beam 151 (in FIG. 3) is then collected by the objective lens 153 and then converted by the optical sensor 154 into an electronic signal. The electronic signals are then digitized by an analog-to-digital converter (ADC) 155 and transmitted to electronic controller 156 for signal processing. The electronic controller may be any electronic processor with suitable processing power, such as a DSP, microcontroller unit (MCU), field programmable gate array (FPGA) or even central processing unit (CPU). In one embodiment, the DSP based controller 156 monitors the electronic signal and then, when the desired component 160 is detected, the two piezoelectric actuator assemblies (each of the piezoelectric actuator assemblies 109, 110, 209, 210) At least one actuator driver (ie, 157a or 157b) may be triggered to drive one of the portions 109, 110 or 219, 220). In another embodiment, the FPGA based controller monitors the electronic signal and then, when the desired component 160 is detected, the two piezoelectric actuator assemblies (each of the piezoelectric actuator assemblies 109, 110, 209, 210) Acts to communicate with the DSP controller to drive one of the parts 109, 110 or 219, 220), or to independently trigger at least one actuator driver (ie 157a or 157b) Do.

  That is, in one embodiment, the plurality of selected or desired components 160 in the channel 164 C of the interrogation chamber 129 is one of the jet channels 127, 128 depending on the desired outflow channels 140, 142 for the plurality of selected components 160. Separated by a jet or buffer or sheath fluid 163 from one. In an exemplary embodiment, the electronic signal activates the driver and triggers the external laminated piezoelectric actuator 219 when multiple target or selected components 160 reach the cross-sectional point of the jet channels 127, 128 and the main channel 164C. (Or activate the driver 157 and trigger the actuator 109). This causes the outer laminated piezoelectric actuator assembly 209 (or 109) to contact and push the diaphragm 170 to compress the jet chamber 130 and a strong jet of buffer or sheath fluid 163 from the jet chamber 130 through the jet channel 127. And push it into the main channel 164 C, thereby pushing the selected or desired component 160 into the outflow channel 142. Note that, similar to the performance of the laminated external piezoelectric actuator assembly 209, triggering of the piezoelectric actuator assembly 210 (or 110) will push the desired component 160 into the outflow channel 140 opposite the jet 128 .

  That is, the sheath or buffer fluid 163 emanating from one of the jet channels 127, 128 has a plurality of target or selection components 160 selected from the respective normal paths of the channel 164C or desired outflow channels 140, 142. Sample diverted toward one of these to separate these multiple target components 160, concentrate the flow of these outlet channels 140, 142, and keep going straight through the outlet channel 141 with the unselected components, if any. The flow of fluid 120 is reduced sharply. That is, the lack of triggering of the piezoelectric actuator assembly 109, 110 means that the plurality of unselected components 160 of the fluid mixture 120 continue to go straight through the outflow channel 141.

  In one embodiment, the plurality of separated components 160 may be stored for storage, for further separation, or for processing such as cryopreservation from one of the first outlet 111 or the third outlet 113. In order to be collected using methods known in the art. Of course, a plurality of components 160 not separated into outlets 111, 113 may also be collected from the second outlet 112. Portions of the first, second and third effluents 111-113 may be characterized electronically to detect component concentrations, pH measurements, cell counts, electrolyte concentrations, and the like.

  In one embodiment, interrogation of sample 120 comprising multiple components 160 (ie, biological material) is accomplished by other methods. That is, a portion or portions of the microfluidic chip 100 may be inspected optically or visually. In general, the plurality of interrogation methods may include direct visual image analysis with a camera or the like, and may utilize direct high intensity light image analysis or fluorescence image analysis. Alternatively, more sophisticated techniques such as spectroscopy, transmission spectroscopy, spectral image analysis, or scattering such as dynamic light scattering or diffuse wave spectroscopy may be used.

  In some cases, light interrogation region 129 binds to or affects multiple components of sample mixture 120, or multiple chemicals that bind to them, or in the presence of a particular material or disease, and / or It may be used in conjunction with multiple additives, such as multiple beads that are functionalized to fluoresce. These techniques may be used to measure cell concentrations to detect diseases or to detect other parameters characterizing components 160.

  However, in other embodiments, multiple polarization backscattering methods may also be used if fluorescence is not used. Using multiple spectroscopic methods, multiple components 160 are interrogated as described above. The spectra of those components 160 that produced significant results and fluoresced (ie, those components 160 that showed a response by the label) were identified for separation by activation of the piezoelectric assemblies 109, 110, 209, 210. Be done.

  In one embodiment, the plurality of components 160 are related to the natural fluorescence or plurality of components 160 of the plurality of components 160 based on the reaction or binding of the components to the plurality of additives or sheaths or buffer fluid 163 It may be specified by using the fluorescence of the substance as a specific tag or background tag, or by fulfilling a selected size, dimension or surface feature etc chosen for the separation.

  In one embodiment, selection of components 160 to be discarded and collected via computer 182 and / or an operator (monitoring electronic signals and triggering piezoelectric assemblies 109, 110, 209, 210) when the assay is complete. May be done.

  In one embodiment, the user interface of computer system 182 includes a computer screen displaying a plurality of components 160 in a field of view for microfluidic chip 100 obtained by CCD camera 183.

  In one embodiment, the computer 182 is used to pump any sample fluid 120, sheath or buffer fluid 163 into the microfluidic chip 100 with an external device such as a plurality of pumps (ie, the pumping mechanism of FIG. 9). In this case, all of these are controlled to further control any heating device that sets the temperature of the fluid 120, 163 introduced to the microfluidic chip 100.

[Chip cassette and holder]
The microfluidic chip 100 is loaded into a chip cassette 212 mounted on a chip holder 200. The tip holder 200 is mounted on a translation stage (not shown) to enable fine positioning of the holder 200. The microfluidic chip holder 200 is configured to hold the microfluidic chip 100 at a position where the light beam 148 may interfere with the plurality of components 160 at the aperture 150 in the manner described above. When the microfluidic chip 100 is in the closed position, the gasket layer 105 (see FIG. 1) forms a substantially leak-free seal between the body 211 and the microfluidic chip 100.

  As shown in FIG. 6, in one embodiment, the microfluidic chip holder 200 is formed of a suitable material, such as an aluminum alloy or other suitable metal / polymer material, and is laminated to the body 211 with at least one And external piezoelectric actuators 209, 210.

  The body 211 of the holder 200 may be of any suitable shape, but the configuration differs depending on the layout of the chip 100. For example, the stacked external piezoelectric actuators 209, 210 must be disposed on the diaphragms 170, 171 such that a contact is formed between the tips of the piezoelectric actuators 219, 220 and the diaphragms 170, 171 of the microfluidic chip 100. It does not. The body 211 of the holder 200 is configured to receive and engage an outer tube (see FIG. 9) to place multiple fluids / samples in fluid communication with the microfluidic chip 100.

  The details of the cassette 212 and the holder 200 and the mechanism for attaching the chip 100 to the cassette 212 and the holder 200 will not be described in any detail, but those skilled in the art are familiar with these devices, It will be appreciated that any configuration may be taken to accommodate the microfluidic chip 100 as long as the multiple objects of the invention are met.

  As shown in FIG. 9, in one embodiment, the pumping mechanism applies pressure to pump the sample fluid mixture 120 from the reservoir 233 (i.e., the sample tube) to the sample input 106 of the tip 100. Including a system having a gas 235.

  A collapsible container 237 having a sheath or buffer fluid 163 therein is disposed in the pressurization tube 236, and the pressurized gas 235 is delivered to the sheath or buffer inlet 107, 108 of the tip 100 by means of the fluid 163 via the tubes 231a and 231b. The fluid 163 is pushed into a manifold 238 having a plurality of different outlets as it is supplied to the

  The pressure regulator 234 regulates the pressure of the gas 235 in the reservoir 233, and the pressure regulator 239 regulates the pressure of the gas 235 in the pipe 236. The mass flow regulators 232a, 232b control the fluid 163 to be pumped to the sheath or buffer inputs 107, 108 respectively via tubes 231a, 231b. That is, the tubes 230, 231a, 231b are used to initially fill the chip 100 with a plurality of fluids 120, and the sample fluid 120 is filled into the sample loading section 106 or the sheath or buffer loading sections 107, 108 throughout the tip 100. May be used to Further, in one embodiment, a tube (not shown) may provide fluid 163 from the manifold 238 to the vents 121, 122, for example, to fill the chambers 130, 131.

  According to an exemplary embodiment, any of the operations, steps, control options, etc. may be implemented by instructions stored on a computer readable medium, such as computer memory, database, etc. Good. Execution of the plurality of instructions stored on the computer readable medium causes the plurality of instructions to cause the computing device to perform any of the plurality of operations, phases, control options, etc. described herein. May be

  The operations described herein are performed by processing circuitry on data received from data processing devices or other sources stored in one or more computer readable storage devices. It may be implemented as multiple actions. A computer program (also known as a program, software, software application, script or code) can be written in any form of programming language including compiled or interpreted language, declarative or procedural language, which is stand-alone It may be arranged in any form including as a program or as a module, component, subroutine, object or other unit suitable for use in a computing environment. A computer program may, but need not, correspond to files in the file system. The program may be part of a file that holds other programs or data (eg, one or more scripts stored in a markup language document), a single file dedicated to the program in question, or It may be stored in a plurality of linked files (e.g., one or more modules, subprograms, or a plurality of files storing portions of code). The computer program may be arranged to be executed by a single computer, or by a plurality of computers interconnected by a communication network, distributed at a single site location or a plurality of sites. The plurality of processing circuits suitable for the execution of a computer program include, by way of example, both general and special purpose microprocessors, and any one or more processors of any kind of digital computer.

  It should be noted that the orientations of the various elements may differ according to other exemplary embodiments, and such variations are intended to be encompassed by the present disclosure.

  The multiple structures and multiple configurations of the microfluidic chip are for illustration purposes only, as shown in various exemplary embodiments. Although only a few embodiments are described in detail in the present disclosure, many modifications (e.g., various modifications) may be made without substantially departing from the novel teachings and advantages of the subject matter described herein. Element sizes, dimensions, structures, shapes and proportions, parameter values, mounting configurations, material applications, color, orientation, etc. variations are possible. Some elements shown as integrally formed may be comprised of multiple parts or elements, and the positions of the multiple elements may be opposite or otherwise different. Well, the nature or number of individual elements or positions may be varied or different. The order or order of steps of any process, logic algorithm or method may be different or may be changed in order according to several alternative embodiments. Other permutations, modifications, alterations and omissions may also be made in the design, operating conditions and configuration of the various exemplary embodiments without departing from the scope of the present disclosure.

Claims (46)

A microfluidic chip comprising a plurality of layers in which a plurality of channels are arranged, wherein
The plurality of channels are
A sample input channel into which a sample fluid mixture of components to be separated is input;
Only two sets of multiple sheath fluid channels,
The plurality of sheath fluid channels of the first set is a first plurality of sheath fluid channels into which a plurality of sheath fluids are injected, the first plurality of sheath fluid channels being the sample at the first intersection point The transverse intersection with the inlet channel allows the plurality of sheath fluids to compress the sample fluid mixture on at least two sides, thereby maintaining the laminar flow in the sample inlet channel, thereby causing the sample fluid mixture to , A relatively smaller thin flow defined by the plurality of sheath fluids,
The second set of sheath fluid channels is a second plurality of sheath fluid channels into which a plurality of sheath fluids are injected, the second plurality of sheath fluid channels being relative to the sample injection channel In the second direction, which is substantially only from above 90 degrees, vertically crossing the sample introduction channel at a second intersection point downstream of the first intersection point, the sample introduction channel A plurality of sheath fluids from the second plurality of sheath fluid channels compress the sample fluid mixture while further maintaining laminar flow, and the plurality of components of the sample fluid mixture are vertically compressed, predetermined. Oriented in the
A plurality of sheath fluid channels of only the two sets;
A plurality of outlet channels extending from the sample input channel, wherein the plurality of components and the plurality of sheath fluids are removed from the microfluidic chip;
Including, microfluidic chips. The plurality of layers have through holes that intersect the sample input channel downstream of the first intersection point and the second intersection point,
The apparatus further comprises a first cover and a second cover disposed at both ends of the through hole to seal the sample introduction channel.
The microfluidic chip according to claim 1 . In interrogation chamber disposed downstream of the second intersection point, the plurality of components of the sample fluid mixture of the sample turned channel, further comprises the interrogation device to interrogate and identify, according to claim 1 or 2. The microfluidic chip according to 2. Displacing the trajectory of the flow of the sample fluid mixture in the sample input channel and extruding a plurality of selected components in the flow of the displaced sample fluid mixture from the interrogation chamber into one of the plurality of outlet channels extending 4. The microfluidic chip of claim 3 , further comprising: a separation mechanism that separates the plurality of selected components from the plurality of components of the sample fluid mixture downstream of the interrogation chamber. At least one jet chamber, the at least one jet chamber including a plurality of sheath fluids introduced into the jet chamber by at least one vent;
At least one jet channel connected to the at least one jet chamber, the at least one jet channel entering the sample input channel in the interrogation chamber;
The microfluidic chip according to claim 4 , further comprising The microfluidic chip according to claim 5 , wherein the separation mechanism comprises at least one piezoelectric actuator assembly disposed on at least one side of the sample injection channel. The microfluidic chip according to claim 6 , wherein the piezoelectric actuator assembly is an external stacked piezoelectric actuator assembly. Further comprising a diaphragm covering each of the jet chambers;
The outer laminated piezoelectric actuator assembly drives the plurality of sheath fluids in the jet chamber into the sample input channel, and the trajectory of the flow of the sample fluid mixture in the sample input channel into the plurality of outlet channels The microfluidic chip according to claim 7 , wherein the diaphragm is aligned with and displaced in order to displace in one of the two. The microfluidic chip according to claim 8 , wherein the external laminated piezoelectric actuator assembly is disposed in a microfluidic chip holder. 9. The microfluidic device of claim 8 , further comprising an electronic circuit connected to the piezoelectric actuator assembly, wherein the electronic circuit amplifies an electronic signal generated by a resistive force from the piezoelectric actuator assembly in contact with the diaphragm. Chip. 11. The microfluidic chip of claim 10 , wherein the electrical signal from the piezoelectric actuator assembly is indicative of an amount of strain generated by the external laminated piezoelectric actuator assembly. 11. The microfluidic chip of claim 10 , wherein an indicator of contact is automatically turned on when contact occurs between the piezoelectric actuator assembly and the diaphragm. 13. The microfluidic chip of claim 12 , wherein the electronic signal activates the indicator if a set threshold is exceeded. 14. The microfluidic chip of claim 13 , wherein the indicator of contact comprises light, sound, touch or any combination thereof. The piezoelectric actuator assembly is
A flexible diaphragm covering the jet chamber;
A piezoelectric material adhered to the top surface of the diaphragm by an adhesion mechanism;
The microfluidic chip according to claim 6 , comprising When voltage is applied to the plurality of electrodes of the piezoelectric actuator assembly, the flexible diaphragm flexes into the jet chamber to deflect the plurality of selected components into one of the plurality of outlet channels; 16. The microfluidic chip of claim 15 , wherein a plurality of sheath fluids are pushed out of the jet chamber into the sample input channel. 6. The microfluidic chip of claim 5 , wherein the jet channel is tapered when connected to the sample input channel. Further comprising microfluidic chip according to any one of claims 1 to 17 a plurality of outlet portions to be arranged in a plurality of end portions of the plurality of outflow channels. 19. The microfluidic chip of claim 18 , wherein the plurality of outlet channels increase in size than the sample inlet channel. 20. The microfluidic chip of claim 19 , further comprising a plurality of notches disposed at the lower end of the microfluidic chip to separate the plurality of outlets. 4. The microfluidic of claim 3 , wherein the sample input channel and the first plurality of sheath fluid channels and the second plurality of sheath fluid channels are disposed in one or more planes of the microfluidic chip. Chip. The sample loading channel and the first plurality of sheath fluid channels and the second plurality of sheath fluid channels are disposed in or between one or more structural layers of the microfluidic chip The microfluidic chip according to claim 3 . 22. The microfluidic device of claim 21 , wherein at least one of the first plurality of sheath fluid channels and the second plurality of sheath fluid channels are disposed in a plane different from the plane in which the sample input channel is disposed. Chip. Wherein at least one of the first plurality of sheath fluid channels and the second plurality of sheath fluid channels are arranged in the structural layer structure different layers the sample turned channel is arranged, according to claim 22 Microfluidic chip. The sample input channel tapers at an entry point to the first intersection with the first plurality of sheath fluid channels and to the second intersection with the second plurality of sheath fluid channels The microfluidic chip according to any one of claims 1 to 24 . 4. The microfluidic chip of claim 3 , wherein the sample input channel is tapered towards the interrogation chamber. The first plurality of sheath fluid channels at a plurality of entry points to the sample input channel at the first intersection point, and the first at a plurality of entry points to the sample input channel at the second intersection point At least one of the second plurality of sheath fluid channel tapers, microfluidic chip according to any one of claims 1 26. The interrogation chamber includes an opening cut through the plurality of structural layers of the microfluidic chip,
The upper window receives a first cover at an opening of at least one of the plurality of structural layers,
23. The microfluidic chip of claim 22 , wherein the lower window receives a second cover at an opening of at least one of the plurality of structural layers. The interrogation chamber includes an opening that is cut through the plurality of planes of the microfluidic chip,
An upper window receives a first cover at an opening of at least one of the plurality of planes of the microfluidic chip,
22. The microfluidic chip of claim 21 , wherein the lower window receives a second cover at an opening of at least one of the plurality of planar surfaces of the microfluidic chip. The interrogation apparatus includes a light source emitting a beam through a first cover to illuminate and excite the plurality of components of the sample fluid mixture,
The microfluidic chip according to claim 3 , wherein the light induced and emitted by the beam passes through a second cover and is received by an objective lens. The interrogation apparatus includes a light source that emits a beam through a plurality of structural layers of the microfluidic chip to illuminate and excite the plurality of components of the sample fluid mixture,
29. The microfluidic chip of claim 28 , wherein the light induced and emitted by the beam is received by an objective lens. The interrogation apparatus includes a light source that emits a beam through the plurality of planes of the microfluidic chip to illuminate and excite the plurality of components of the sample fluid mixture;
30. The microfluidic chip of claim 29 , wherein the light induced and emitted by the beam is received by an objective lens. Displacing the trajectory of the flow of the sample fluid mixture in the sample input channel and extruding a plurality of selected components in the flow of the displaced sample fluid mixture from the interrogation chamber into one of the plurality of outlet channels extending Further comprising: a separation mechanism for separating the plurality of selected components from the plurality of components of the sample fluid mixture downstream of the interrogation chamber,
The separation mechanism includes at least one piezoelectric actuator assembly disposed on at least one side of the sample injection channel,
32. The microfluidic chip of claim 31 , wherein the emitted light received by the objective lens is converted into an electronic signal that triggers the piezoelectric actuator assembly. Displacing the trajectory of the flow of the sample fluid mixture in the sample input channel and extruding a plurality of selected components in the flow of the displaced sample fluid mixture from the interrogation chamber into one of the plurality of outlet channels extending Further comprising: a separation mechanism for separating the plurality of selected components from the plurality of components of the sample fluid mixture downstream of the interrogation chamber,
The separation mechanism includes at least one piezoelectric actuator assembly disposed on at least one side of the sample injection channel,
33. The microfluidic chip of claim 32 , wherein the emitted light received by the objective lens is converted into an electronic signal that triggers the piezoelectric actuator assembly. Wherein one of the sample fluid mixture or said plurality of sheath fluid, the pumping device, said pumped into the microfluidic chip, microfluidic chip according to any one of claims 1 34. External tube, a plurality of fluid, the microfluidic chip to communicate with the micro-fluidic chip according to any one of claims 1 35. The microfluidic chip according to any one of claims 1 to 36 , wherein the plurality of components are a plurality of cells. The plurality of cells to be separated may be alive from non-viable or non-viable sperm or spermatozoa separated by sex and other sex classification variations, a plurality of stem cells separated from a population cell, a plurality of sperm cells one or more labeled cells, a plurality of genes has been separated in the nuclear DNA of the characteristics of the particular, multiple cells isolated based on the plurality of surface markers that are separated from a plurality of unlabeled cells containing, film Multiple cells separated based on integrity or viability, multiple cells separated based on possible or predicted regeneration status, multiple cells separated based on frozen viability, multiple contaminants Or multiple cells separated from debris, multiple healthy cells separated from multiple damaged cells, multiple white blood cells and platelets of plasma mixture The number of red blood cells, or at least one of a plurality of any cell which is divided into plural corresponding fragment from a plurality of any other cell components, the microfluidic chip according to claim 37. The system according to claim 1, wherein the plurality of components to be separated are moved to one of the plurality of outlet channels, and the plurality of unselected components flow out through the other of the plurality of outlet channels. The microfluidic chip according to any one of 38 . 36. The microfluidic chip of claim 35 , further comprising a computer that controls the pumping of the sample fluid mixture or the plurality of sheath fluids to the one of the microfluidic chips. 29. The microfluidic chip of claim 28 , further comprising a computer displaying the plurality of components in a field of view acquired by a CCD camera disposed over the opening of the microfluidic chip. 30. The microfluidic chip of claim 29 , further comprising a computer that displays the plurality of components in a field of view acquired by a CCD camera disposed over the opening of the microfluidic chip. A microfluidic chip loaded in a microfluidic chip cassette mounted on a microfluidic chip holder, the sample loading unit for introducing a sample fluid containing a plurality of components into the microfluidic chip; A plurality of sheath inlets for introduction to a fluidic tip and a sample channel for flowing the sample fluid, wherein the sample channel is connected to only two sets of sheath fluid channels for flowing the sheath fluid downstream, the sample channel being At a point of intersection of the first set of sheath fluid channels in a first direction, the sample channel being substantially only from above 90 degrees with respect to the sample channel at a second point of intersection; The sample channel being connected to the second set of sheath fluid channels in two directions; A microfluidic chip that,
The sample fluid is pumped from a reservoir to the sample loading portion of the microfluidic chip, and the sheath fluid is pumped to the plurality of sheath loading portions of the microfluidic chip, the sample fluid and the sheath fluid being the sample A pumping mechanism for pumping downstream from the inlet and the plurality of sheath inlets through the sample channel;
, A microfluidic chip system. The microfluidic chip has a through hole that intersects the sample channel downstream of the first intersection point and the second intersection point, and is disposed at both ends of the through hole to seal the sample channel. 44. The microfluidic chip system of claim 43 , further comprising one cover and a second cover. Injecting a sample fluid mixture comprising a plurality of components into a sample injection channel of the microfluidic chip;
Injecting a plurality of sheath fluids into only two sets of sheath fluid channels;
The plurality of sheath fluids enter a plurality of sheath fluid channels of a first set of the microfluidic chip, and the plurality of sheath fluids from a plurality of sheath fluid channels of the first set are of the first set The plurality of sheath fluid channels and the plurality of sheath fluids from the plurality of sheath fluid channels of the first set merge with the sample fluid mixture of the sample introduction channel at a first intersection of the plurality of sheath fluid channels and the sample input channel. Compressing the sample fluid mixture in one direction of the sample input channel to concentrate the plurality of components of the sample fluid mixture around the center of the sample input channel;
The plurality of sheath fluids enter a plurality of sheath fluid channels of the second set of microfluidic chips, and the plurality of sheath fluids from the plurality of sheath fluid channels of the second set are at the first intersection point. The second set of sheaths of the second set merges with the sample fluid mixture of the sample input channel at a second intersection of the second set of sheath fluid channels and the sample input channel downstream. The plurality of sheath fluids from the fluid channel are such that the plurality of components are concentrated and aligned in the sample input channel by both width and depth as the components flow through the sample input channel. , the sample fluid mixture, over substantially 90 degrees with respect to the sample charged channel at the second intersection Further compressed in a second direction which is only from said plurality of sheath fluid, when the plurality of components to flow through the sample turned channel, to compress and directing the plurality of components in the selected direction, the Act on multiple components,
A method of orientating multiple components of a fluid mixture. The microfluidic chip has a through hole that intersects the sample injection channel downstream of the first intersection point and the second intersection point, and the sample injection channel is sealed at both ends of the through hole. A first cover and a second cover are arranged,
46. A method of directing components of a fluid mixture according to claim 45 .

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