JP2015047133A - Device and method for separating substances having different densities - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a device and a method for separating substances having different densities.SOLUTION: There is provided the device for separating substances having different densities. A cup body has an internal cavity configured to hold a medium. An inner wall defines a center body region having a top edge and a bottom edge. The top edge is wider than the bottom edge. An internal shoulder circumscribes the top edge of the center body region. The internal shoulder defines a neck region above the center body region, and defines a shoulder trap below the neck region. The shoulder trap circumscribes the top edge of the center body region, and is wider than the neck region. When the device is caused to spin about a central axis, the medium moves upward along the inner wall and toward the shoulder trap.

Description

(Technical field)
The present invention relates to automated and semi-automated electrophysiological analysis, and in particular to high throughput automated electrophysiology. To this end, a separation device for receiving a medium, an apparatus having a separation cup, and a separation method are provided.

(background)
The electrical behavior of cells and cell membranes is important in basic research and in modern drug development. A particular area of interest in this area is in ion channel and transporter research. Ion channels are pores based on proteins found in the cell membrane that are responsible for maintaining an electrochemical gradient between the extracellular environment and the cytoplasm of the cell. An ion channel is a passive element in that once opened, it flows in the direction of the electrochemical gradient in which the ions are present.

  Ion channel research is a very diverse and fruitful area that encompasses basic academic research, as well as biotechnological and pharmaceutical research. Electrophysiology is performed on isolated cell membranes or vesicles, as well as on synthetic membranes in which solubilized channels are reconstituted into the manufactured membrane. Instruments for automated, high-throughput studies of ion channels have been developed and can be referred to as high-throughput electrophysiological measurement systems.

  In general, many types of assays require sufficient cell preparation. Accordingly, a need continues to exist for effective methods for cell preparation, and apparatus or devices configured to perform such methods. Sufficient cell preparation is of particular interest in terms of automated assays, which can perform multiple assays with minimal human intervention and increase throughput, thus gaining per day The number of data points that are played can be increased. Known automated high-throughput measurement systems (eg, those described above) can be employed to perform electrophysiological assays in a relatively quick and efficient manner, but such systems are It is not equipped to automatically prepare cells and cell solutions used in the assay.

  It is an object of the present invention to provide a convenient device, apparatus and method for automatically preparing cells and cell solutions for use in electrophysiological assays.

The present invention provides, for example:
(Item 1)
A device (300) for separating materials of different density, the device comprising:
A cup body (304) having an internal cavity (302) configured to hold a medium;
An inner wall (306) defining a central body region (308) having an upper end and a lower end, the upper end being wider than the lower end, an inner wall (306); and circumscribing the upper end of the central body region (308) An internal shoulder (318), wherein the internal shoulder defines a neck region (310) above the central body region and a shoulder trap (320) below the neck region, the shoulder trap An inner shoulder (318) circumscribing the upper end of the central body region
Comprising:
The shoulder trap (320) is wider than the neck region (310) so that when the device is spun around a central axis, the media is directed upward along the inner wall (306) and the shoulder Moving toward the trap (320), a relatively high density material in the medium is collected in the shoulder trap, and a relatively low density material in the medium is an opening above the neck region. Device (300) ejected from the device through section (322).
(Item 2)
The inner cavity (302) in the central body region (308) has a substantially conical shape, so that the diameter of the inner cavity (302) in the central body region is the same as that of the central body region. The device according to the above item, which is tapered downward toward the lower end.
(Item 3)
Any of the preceding items comprising a coupling member (328) configured to couple the device to a rotary actuator (206), wherein the cup body (304) is rotatable about a central axis of the cup body. A device according to
(Item 4)
The device according to any of the preceding items, wherein the inner wall (306) defines a bottom region below the central body region (308), and the bottom region has a substantially hemispherical shape.
(Item 5)
Above the neck region (310) is further provided an opening (324) defining the opening (322), through which the relatively low density material is discharged, the opening being A device according to any of the preceding items, circumscribing the neck region and extending outward in a direction away from the neck region.
(Item 6)
And further comprising one or more passages (602) formed through the cup body (304) from the lower end of the central body region (308) to the upper end of the central body region, each of the one or more passages , Defining a flow path from the lower end to the upper end of the central body region, and in particular, each of the one or more passageways (602) is a longitudinal bore (604) formed through the cup body (304). The longitudinal bore includes a lower aperture formed in the inner wall (306) near the lower end of the central body region (308) and a shoulder trap near the upper end of the central body region. The device according to any of the preceding items, connected to an upper aperture formed in the inner wall below (320).
(Item 7)
A method of separating a cell suspension from a starting mixture, the starting mixture comprising a relatively low density buffer solution and a relatively high density of cells, the method comprising:
Dispensing the starting mixture to a device (204, 300, 600), the device comprising a cup body (304) having an internal cavity (302) for holding the starting mixture; A device, in particular, is a device according to any of the above items;
Spinning the device about a central axis so that the starting mixture is directed upward along the inner wall (306) of the cup body (304) and circumscribing the inner cavity (320) ) So that the relatively high density cells in the starting mixture are collected in the shoulder trap and at least some of the relatively low density buffer solution in the starting mixture is Evacuating from the device through an opening (322) in the cup body;
Dispensing a desired solution into the internal cavity (302) of the cup body (304) after draining at least some of the relatively low density buffer solution; and the desired solution and the relatively high Stirring the device such that the cells of density are mixed together.
(Item 8)
Stirring the device (300) includes:
Accelerating the device forward at a predetermined forward acceleration rate;
Maintaining a predetermined forward spin rate for a first predetermined amount of time;
Decelerating the device at a predetermined forward deceleration rate;
Accelerating the device backwards at a predetermined backward acceleration rate;
Maintaining a predetermined backward spin rate for a second predetermined amount of time;
Decelerating the device at a predetermined backward deceleration rate; and at least one of pulverizing or titrating the desired solution and the relatively dense cells with a pipettor. Method.
(Item 9)
An apparatus (200) for separating materials of different densities, the apparatus comprising:
A separation cup (204, 300), in particular a separation device according to any of the above items, wherein the separation cup has an internal cavity (302) configured to hold a medium; And a shoulder trap (320) circumscribing the internal cavity, wherein the shoulder trap causes the relatively high density material in the medium to move to the shoulder trap (when the separation cup is spun around a central axis). 320) and configured to drain relatively low density material in the medium from the separation cup (204, 300); and the separation cup A rotary actuator (206) coupled to the rotary actuator, the rotary actuator being configured to spin the separation cup about a central axis of the separation cup Eta (206)
An apparatus (200) comprising:
(Item 10)
The separation cup (204, 300) is:
An inner wall (306) defining a central body region (308) having an upper end and a lower end, the upper end being wider than the lower end, an inner wall (306); and circumscribing the upper end of the central body region (308) An inner shoulder (318), the inner shoulder further comprising an inner shoulder defining the shoulder trap (320) circumscribing the inner cavity (302) and a neck region above the central body region;
In particular, the internal cavity (302) in the central body region (308) of the separation cup has a substantially conical shape so that the diameter of the internal cavity in the central body region is equal to the central body. An apparatus according to any of the preceding items, wherein the apparatus is tapered downward toward the lower end of the region.
(Item 11)
The diameter of the internal cavity (302) of the separation cup (204, 300) increases downward through the shoulder trap (320), and the diameter of the internal cavity of the separation cup is determined by the neck And / or the separation cup (204, 300) further comprises an opening (324) circumscribing the neck region above the neck region (310). And the opening (324) extends outwardly away from the neck region,
An apparatus according to any of the above items.
(Item 12)
The separation cup (204, 300, 600) further includes one or more passages (602) formed through the separation cup from the lower end of the central body region (308) to the upper end of the central body region. Each of the one or more passageways (602) defines a flow path from the lower end to the upper end of the central body region of the separation cup;
An apparatus according to any of the above items.
(Item 13)
Each of the one or more passageways (602) of the separation cup (204, 300, 600) comprises a longitudinal bore (604), the longitudinal bore being in the central body region (308). A lower aperture (606) formed in the inner wall (306) near the lower end and an upper aperture (608) formed in the inner wall near the upper end of the central body region and below the shoulder trap (320). The device according to any of the above items, which is connected to the device.
(Item 14)
A housing (400) surrounding the separation cup (204, 300, 600), the housing comprising a collection chamber (412), circumscribing the opening (324) of the separation cup; As a result, when the separation cup is spun around the central axis, the collection chamber receives the relatively low density material discharged from the separation cup, and in particular the collection chamber (412) Floor (414) is inclined towards the drain (416, 418) so that the relatively low density material discharged from the separation cup can be drained from the collection chamber. A device according to any of the preceding items.
(Item 15)
Further comprising a mounting assembly (220, 500) for coupling said separating cup (204, 300, 600) to said rotary actuator (206), said separating cup being removably fixable within said mounting assembly; A device according to any of the items.

(Summary)
Devices (204, 300, 600) are provided for separating materials of different densities. The cup body (304) has an internal cavity (302) configured to hold the media. Inner wall (306) defines a central body region (308) having an upper end and a lower end. This upper end is wider than this lower end. The inner shoulder (318) circumscribes the upper end of this central body region (308). The inner shoulder defines a neck region (310) above the central body region (308) and a shoulder trap (320) below the neck region. The shoulder trap (320) circumscribes the upper end of the central body region (308) and is wider than the neck region (310). As the device is spun around the central axis, the media moves upward along the inner wall (306) towards the shoulder trap (320). The relatively high density material in the media is collected in the shoulder trap and the relatively low density material is exhausted from the device through the opening (322) above the neck region (310). . Furthermore, the present invention provides an apparatus (200) for separating media and a method for separating media using this separation device.

(Summary)
The invention is defined in claims 1, 7 and 9, respectively. Specific embodiments are set forth in the dependent claims.

  A device is provided for separating materials of different densities. The cup body has an internal cavity configured to hold the media. The inner wall defines a central body region having an upper end and a lower end. This upper end is wider than this lower end. The inner shoulder circumscribes the upper end of this central body region. The inner shoulder defines a neck region above the central body region and a shoulder trap below the neck region. The shoulder trap circumscribes the upper end of the central body region and is wider than the neck region. As the device is spun around the central axis, the media moves upward along the inner wall toward the shoulder trap. The relatively high density material in the medium is collected in the shoulder trap and the relatively low density material is exhausted from the device through an opening above the neck region.

  In one embodiment, the separation device is designed according to details about the separation cup as specified in the apparatus claims or as specified in the detailed description. The features disclosed herein may be combined individually or in any arbitrary combination.

  A method of separating a cell suspension from a starting mixture comprising a relatively low density buffer solution and a relatively high density of cells is also provided. The starting mixture is dispensed into a device, the device having a cup body having an internal cavity for holding the starting mixture. The device is spun around the central axis so that the starting mixture is driven upward along the inner wall of the cup body towards the shoulder trap circumscribing the inner cavity. The relatively high density of cells in the starting mixture is collected in the shoulder trap, and at least some of the relatively low density buffer solution in the starting mixture is removed from the device through the opening in the cup body. Discharged.

  According to a preferred embodiment of the method, the spinning step comprises flowing at least some of the starting mixture through one or more passages of the cup body so that a relatively high density of material can be obtained. Deposits in this shoulder trap as it exits one or more passages. Preferably, these passages are designed as specified in the device claims or as specified in the detailed description.

  Further provided is an apparatus for separating different density materials. The separation cup has an internal cavity configured to hold the media. The separation cup includes a shoulder trap that circumscribes the internal cavity. The shoulder trap collects relatively high density material in the medium when the separation cup is spun around the central axis, and relatively low density material in the medium. Is configured to be discharged from the separation cup. A rotary actuator is coupled to the separation cup and configured to spin the separation cup about the central axis of the separation cup.

  Preferably, the separation cup of the apparatus is designed according to the device described above, for the separation described in the claims, or according to the features specified in the detailed description. These features can be combined individually or in any arbitrary combination.

  The invention can be better understood with reference to the following drawings. The components in the drawings are not necessarily to scale, emphasis instead being placed upon illustrating the principles of the invention. In the drawings, like reference numerals designate corresponding parts throughout the different views.

FIG. 1 is a plan view of an example embodiment of an automated high-throughput electrophysiological measurement system. FIG. 2 is a side view of an example embodiment of a robotic pipettor head of an automated high throughput electrophysiological measurement system. FIG. 3 is a cross-sectional side view of an exemplary embodiment of a patch plate supported on a plenum of an automated high throughput electrophysiological measurement system. FIG. 4A is an exploded view of an example of one embodiment of an apparatus for separating materials of different densities. FIG. 4B is a front upper right perspective view of the automated device for separating materials of different densities of FIG. 4A shown in an assembled configuration. FIG. 5 is a top perspective view of an example of one embodiment of a separation cup. 6 is a side sectional view of the separation cup of FIG. FIG. 7 is a top perspective view of an example of one embodiment of a cup housing. 8 is a side sectional view of the cup housing of FIG. FIG. 9 is a side cross-sectional view of the top of the automated device for separating different density materials of FIG. 4A shown in an assembled configuration. FIG. 10 is a perspective view of an example of one embodiment of a cup installation assembly. FIG. 11 is a side cross-sectional view of the lower portion of the automated device for separating different density materials of FIG. 4A shown in an assembled configuration. FIG. 12 is a top perspective view of an example of an alternative embodiment of a separation cup. FIG. 13 is a flowchart of exemplary method steps for automatically separating materials of different densities. FIG. 14 is a flowchart of exemplary method steps for automatically agitating a separation cup of an automated apparatus for separating materials of different densities.

(Detailed explanation)
Apparatus, devices, systems and methods for separating different density materials are shown and described. This device can be utilized, for example, for automated preparation of cells for use in biological assays, such as electrophysiological assays performed by automated high-throughput electrophysiological measurement systems. Preparation of cells for biological assays can include, for example, ensuring that cells are suspended in a desired buffer solution at a desired concentration and with a desired level of homogeneity. However, it is understood that this device can be used to separate different densities of material in a way that goes beyond cell preparation for biological assays.

  As disclosed herein, an apparatus (or device, or system) for separating materials of different densities may comprise a separation cup configured to hold a flowable medium, and this flowability The medium can include materials of different respective densities. The separation cup may include one or more internal features that separate different densities of material held in the separation cup. A method for preparing a solution of cells is also disclosed, including a method of suspending cells in a desired buffer solution at a desired concentration and at a desired level of homogeneity. In some embodiments, an apparatus for preparing a substance can be utilized in performing one or more of these methods. For example, the cell controls one or more operating parameters of the device (eg, separation cup acceleration and deceleration rates, separation cup rotation speed, separation cup spin direction, and spin duration), Can be prepared. This device can be provided in the process deck of an automated high-throughput electrophysiological measurement system, which can allow the on-deck preparation of cell concentrates. These and other aspects of cell preparation devices and methods are discussed in further detail below.

  Referring to FIG. 1, an example implementation of an automated high throughput electrophysiological measurement system 100 is shown in plan view. The system 100 in this example is configured to perform simultaneous measurements on multiple samples (eg, a two-dimensional grid or array of samples). The high throughput electrophysiological measurement system may include a measurement platform 102, which is also referred to as a process deck 102. The process deck 102 may support various components of the system 100 and a substantially planar surface to support or maintain a desired spatial arrangement of some or all of the components of the measurement system 100. Can be provided.

  The system may include a control module 104 that controls the operation of the system 100 during the assay. The control module 104 can include, for example, an external microcomputer, a display device, and a software user interface. The control module 104 may also include a microcontroller that is interfaced to an external microcomputer to control real-time functional aspects of the system 100 including motion control, fluidic control, and electrical data recording.

  The system 100 may also include a patch engine that controls the components of the system 100, performs electrophysiological measurements, and digitizes data acquired during the patch clamp assay. In this example, the patch engine includes a plenum 106, an electrode plate 108, and a data acquisition engine. These components are discussed in further detail below.

  The system 100 may include multiple stations or modules configured to perform various functions. In the illustrated example, the system 100 includes seven stations: a tip rack station 110; an external buffer station 112; a first compound station 114; an analysis station 116; a wash station 118; a second compound station 120; 122 is included. It will be appreciated that the system 100 may include more or fewer stations, including stations that provide functions different from those just described.

  In this example, each of these stations is shaped to receive an SBS-standard 384 well microtiter plate (Society for Biomolecular Sciences). In other words, in this example, these stations can be described as having an SBS-standard 384 well microplate footprint. The assay steps take place in the process deck 102 and a robotic pipettor head delivers fluid from the external buffer station 112, cell station 122, and compound stations 114 and 120 to the measurement substrate at the analysis station 116. This robot head is further discussed below with reference to FIG.

  The measurement substrate may be referred to as a patch plate and may include a plurality of holes or holes around which corresponding samples (eg, cells or cell membranes) are positioned or sealed for analysis. In this example, the patch plate is an SBS-standard 384 well microplate. Thus, the patch plate in this example includes 384 individual wells for holding cells, external buffer, and biological screening compounds. In this example, the 384 wells of the patch plate can be arranged in a grid of 16 columns (identified by AP) and 24 columns (1-24). The well of the patch plate can include one or more holes formed through the lower surface. Each hole may have a diameter of about 2 micrometers (μm), for example. A patch plate with one hole per well may be referred to as a single hole plate. A patch plate with multiple holes per well (eg, an array of 64 holes) may be referred to as a collective patch clamp (PPC) plate. The patch plate can be moved to and from the analysis station during the assay. This patch plate is discussed in further detail below with reference to FIG.

  The tip rack station 110 holds a tray on which pipettor tips can be preloaded. The robotic pipettor head is lower towards the chip rack station 110 and loads the pipettor chip at the start of the assay. This pipettor tip can be utilized to aspirate and dispense external buffer solutions, compounds, and cells at the appropriate number of times during a given assay, depending on the particular method indicated for the assay.

  External buffer station 112 may also be referred to as an input station and may include an external buffer boat that holds an external buffer solution. In some example implementations, peristaltic pumps and vacuum assisted waste bottles are selectively employed to automatically fill and drain the external buffer station 112. This external buffer boat can be filled with an external buffer solution before the start of the assay. The external buffer solution can be a physiological salt solution containing a salt or mixture of salts that mimics the extracellular solution (eg, a solution containing a low concentration of potassium). The robotic pipetter head can aspirate the external buffer solution from the external buffer station 112, transport the external buffer solution to the analysis station 116, and dispense the buffer solution into the wells of the patch plate.

  The first and second compound stations 114 and 120 are also referred to as input stations and hold biological screening compounds or other types of reagents that can be utilized during the assay. SBS-standard 384 well compound plates (eg, microplates) can hold biological screening compounds and are within the footprint of the first or second compound station 114 or 120. The robotic pipetter head can similarly aspirate the compound from the compound station 114 or 120, transport the compound to the analysis station 116, and dispense the compound into the wells of the patch plate.

  The analysis station 116 includes a patch engine plenum 106 and supports the patch plate during the assay. The plenum 106 includes a reservoir 126 and an internal buffer solution can be pumped into and out of the plenum reservoir 126 from below during the assay. The internal buffer solution can be a salt solution (eg, a solution containing a high concentration of potassium) that contains a salt or mixture of salts that mimics the internal cytoplasm of viable cells. The patch plate rests on the plenum 106 and an o-ring 128 that surrounds the periphery of the plenum 106 creates a hermetic seal between the patch plate and the plenum reservoir 126. A small negative (differential) pressure is introduced, which pushes the cells (or cell membrane) in the well toward the hole at the bottom of the well. This differential pressure therefore forms a high resistance electrical seal between the cell (or cell membrane) and the bottom of the well, as will be appreciated by those skilled in the art.

  Electrode plate 108 is also referred to as an electronics head and is used to perform electrophysiological measurements on cell samples in a patch plate. Electrophysiological measurements can be performed by creating an electrical circuit across the holes in the wells of the patch plate. An electrical circuit can be formed by positioning the electrodes on opposite sides of the patch plate membrane. For example, the sensing electrode can be positioned above the membrane and the ground electrode can be positioned below the membrane. Thus, in this example, the plenum 106 may include four ground electrodes 130 positioned at the top of the plenum reservoir 126, and in this example, the electrode plate 108 is in a frame that fits the top of the patch plate and the plenum. May include an array of sensing electrodes 132 housed in the housing. The electrode plate 108 is discussed in further detail below with reference to FIGS.

  The array of sensing electrodes 132 on the electrode plate 108 may correspond to the array of wells in the patch plate, and each sensing electrode 132 may perform an electrophysiological measurement on an individual well in the patch plate. Thus, in this example, electrode plate 108 may include an array of 384 sensing electrodes 132. Each sensing electrode 132 corresponds to one electronic channel. Thus, in this example, 384 sensing electrodes 132 correspond to 384 electron channels.

  The electrode 132 can be, for example, silver or a silver coated pin (eg, Ag / AgCl). To complete the circuit, an appropriate electrolyte (eg, salt) solution can be added to the wells of the patch plate and the plenum reservoir 126. For example, the external buffer solution and the internal buffer solution may include chloride ions to allow the sensing electrode 132 and the ground electrode 130 to monitor electrical activity.

  Electrode plate 108 can be clamped to plenum 106 during the assay, and sensing electrodes 132 are received in individual wells of the patch plate. The electrode plate 108 may include holes formed through its upper surface, providing access to the pipettor tip. In this way, the electrode plate 108 allows for the addition of compounds to the patch plate wells while simultaneously measuring the ionic current in the wells. As discussed in more detail below with reference to FIG. 3, sensing electrode 132 and ground electrode 130 may be coupled to measurement electronics to obtain data related to electrophysiological measurements.

  Wash station 118 can receive various components and cleans these components after the assay. In this example, the cleaning station 118 includes a reservoir that contains pipettor tips and electrode pins of the electrode plate for cleaning procedures. Thus, in this example, the wash station 118 may include a manifold of input ports that match the dimensions of the pipettor tips and electrode pins. A fluid handling system (not shown) may pump cleaning solution through the cleaning station 118, which is a large boxed carboy under the process deck 102 (not shown). Poured into. The cleaning station 118 can also serve as a stationary position for the electrode plates when not in use. The robotic pipettor head can pick up the electrode plate 108 at the wash station 118 and transport the electrode plate 108 to the analysis station 116 during the assay. The robotic pipetter head can then return the electrode plate 108 to its rest position at the wash station 118 at the end of the assay.

  Cell station 122 may also be referred to as an input station and includes a cell boat that holds cells (or other biological samples) used in the assay. The cells can be dispersed in an external buffer solution while present in the cell boat. The robotic pipettor head can similarly aspirate cells from the cell station 122, transport the cells to the analysis station 116, and dispense the cells into the wells of the patch plate.

  Referring to FIG. 2, an example implementation of a robotic pipettor head 150 for the high throughput electrophysiological measurement system 100 is shown in side view. The robot pipetter head 150 may also be referred to as a fluidic head or a multi-channel dispensing head. The robotic pipetter head 150 can be used to add, remove, or transfer fluids, cell solutions, and compounds into the wells of a patch plate. In this example, the robotic pipettor head 150 may hold a pipettor chip that is utilized to transport fluid from the cell station 122, the external buffer station 112, and the compound stations 114 and 120 to the analysis station 116. Pipettor tips aspirate or dispense fluid in the correct amount according to the assay protocol.

  The robotic pipetter head 150 may be coupled to a three-dimensional mechanical gantry system 152 for moving the robotic pipettor head 150 between stations in the process deck. The control module 104 can communicate with the mechanical gantry system 152 and controls the movement of the robotic pipetter head 150 during the assay.

  At the start of the assay, the robotic pipettor head 150 can move to the chip rack station 110 and load the pipettor chip. The robot pipettor head 150 is also provided as a transport mechanism for the electrode plate 108. Thus, the robot pipetter head 150 can include, for example, an electrode plate transport clip 154 that holds the electrode plate 108. The robotic pipettor head 150 can load the electrode plate 108 from its rest position at the wash station 118 and transport it to the analysis station 116 where it clamps to the plenum 106 during the assay. At the end of the assay, robotic pipettor head 150 can load electrode plate 108 from analysis station 116 and transport it back to wash station 118.

  Referring now to FIG. 3, an exemplary implementation of the patch plate 160 supported by the plenum 106 of the high throughput electrophysiological measurement system 100 is shown. The patch plate 160 may include a plurality of wells 162 (eg, 384 wells) as discussed above. Two wells 162 of the patch plate 160 are shown for illustrative purposes in FIG. Each well 162 is bounded by a well wall 164 and bounded by a bottom 166 of the patch plate 160. Further, the wells 162 in the exemplary patch plate 160 of FIG. 3 include holes 168 that are each formed through the bottom 166 of the patch plate 160. Cells 170 in individual wells 162 can be sealed to the bottom 166 of patch plate 160 by differential pressure, as discussed above.

  The well 162 of the patch plate 160 can be filled with an external buffer solution 172 and the plenum reservoir 126 of the plenum 106 located under the patch plate 160 can be filled with an internal buffer solution 174. The sensing electrodes 132 can be positioned in the individual wells 162 of the patch plate 160 and, as will be appreciated by those skilled in the art, the electrical that occurs in the wells 162 during the assay, such as the activity of the ion channels 176 of the cells 170. Activity is measured. The ground electrode 130 of the plenum 106 completes an electrical circuit that traverses the individual holes 138 of the patch plate 160.

  The sensing electrode 132 and ground electrode 130 of the electrode plate 108 are routed through measurement electronics 180, such as a programmable voltage source (not shown), an amplifier 182 and an analog-to-digital converter (ADC) 184, for example. And can communicate with the data acquisition engine 178. As seen in the example shown in FIG. 3, sense electrode 132 and ground electrode 130 are in signal communication with amplifier 182, which is in signal communication with ADC 184, which in turn signals with data acquisition engine 178. Communicating. Amplifier 182 may be a high gain, low noise trans-impedance amplifier that converts the current measured on sense electrode 132 into an analog voltage signal. The ADC 184 may convert this analog voltage signal from the amplifier 182 into a digital voltage measurement. The data acquisition engine 178 can therefore record this digital voltage measurement for the sensing electrode channel in the computer memory.

  Referring now to FIG. 4A, an exploded view of an example of one embodiment of an apparatus 200 for separating different density materials is shown. The installation frame 202 supports the separation cup 204 and the rotary actuator 206. The rotary actuator 206 drives the rotation of the separation cup 204 about the central axis to separate different density materials. The installation frame 202 may be part of a process deck 102 of an automated high-throughput electrophysiological measurement system 100, for example, as illustrated in FIGS. As discussed further below, the separation cup 204 allows a relatively low density material to exit the cup 204 through an opening 208 at the top of the cup 204 while capturing a relatively high density material. With internal features.

  The rotary actuator 206 includes a motor 210 and is installed below the installation frame 202 by an actuator housing 212. Separation cup 204 is received within cup housing 214 and is disposed within cup housing 214. The cup housing 214 is installed on the upper side of the installation frame 202 and can be fixed to a pair of support frames 216 (which are also installed on the upper side of the installation frame 202).

  As discussed further below, the separation cup 204 is coupled to the rotary actuator 206 through an opening 218 formed in the installation frame 202 by a cup installation assembly 220 and a rotor assembly 222. The cup mounting assembly 220 includes a cup mount 224, a support pad 226, a biasing spring 228, and a rotary actuator mount 230 in this example. The rotor assembly 222 includes a pair of ball bearings 232 that facilitate rotation of the cup mounting assembly 220 in this example. The rotor assembly 222 may also include one or more washers 234 that may be used to selectively adapt different types of motors to the actuator housing 212.

  As seen in FIG. 4A, the cup housing 214 includes a drain tube 236 in this example, which drains through the drain opening 240 formed through the installation frame 202 and the fluid. Get in touch. The drain nozzle 238 may be connected to a piping system (not shown) for the purpose of removing relatively low density material that is centrifugally removed from the separation cup 204. As also seen in FIG. 4A, a housing cap 242 may cover the top surface of the cup housing 214 for the purpose of confining the separation cup 204 when the separation cup 204 is placed in the cup housing 214. An opening 244 is formed through the housing cap 242 to allow the pipetter to access the interior of the cup housing 214 in this example. The opening 244 of the housing cap 242 is located above the separation cup 204 in this example so that the pipettor can aspirate material from the separation cup 204 or dispense material into the separation cup 204. obtain.

  FIG. 4B is a front upper right perspective view of the apparatus 200 for separating the different density materials of FIG. 4A shown in an assembled view. As seen in FIG. 4B, the separation cup 204 is placed in the cup housing 214, and the cup housing 214 is installed on the installation frame 202. The separation cup 204 is coupled to the rotary actuator 206 through an opening 218 formed through the installation frame 202. Accordingly, the rotary actuator 206 can drive the rotation of the separation cup 204 to discharge a relatively low density material from the separation cup 204. As will be discussed further below, relatively low density material removed from the separation cup 204 by centrifugal force may be drained into the drain tube 236 and through the drain nozzle 238 (as shown). Is not).

  The motor 210 may be a brushless direct current (DC) motor coupled to a corresponding encoder 246. The motor 210 may rotate the installation assembly 220 and the separation cup 204 secured within the installation assembly 220 about a central axis. The motor 210 of the rotary actuator 206 may communicate with a controller (eg, a servo drive controller) that controls the operation of the motor 210 to rotate the installation assembly 220 and the separation cup 204. This controller may be in signal communication with the control module 104 discussed above with reference to FIG. 1 and may be driven by the control module 104 in this example. In this manner, the control module 104 may control one or more operational parameters (eg, acceleration and deceleration of rotation of the separation cup 204, rotation speed, and duration of rotation). The control module 104 may in this example be the control module discussed above with reference to FIG.

  Referring now to FIGS. 5 and 6, a top perspective view and a side cross-sectional view of an example of one embodiment of a separation cup 300 are shown, respectively. The separation cup 300 includes an internal cavity 302 formed in the body 304 (FIG. 6) of the separation cup 300. The inner cavity 302 of the separation cup body 304 can hold a flowable medium (eg, a solution, suspension, slurry, emulsion, etc.). The internal cavity 302 can hold a starting mixture, including, for example, a relatively high density material to be separated and a relatively low density material. This starting mixture can be, for example, a cell suspension comprising cells suspended in a medium (eg, serum, buffer, antibiotics, etc.). Accordingly, the inner wall 306 of the separation cup 300 that at least partially defines the inner cavity 302 is provided with various regions of the separation cup 300 (eg, a central body region 308, a neck region 310, and A bottom region 312) may be further defined. The neck region 310 of the separation cup 300 is located above the central body region 308 in this example, and the bottom region 312 of the separation cup is located below the central body region 308 in this example.

  The contour of the inner wall 306 facilitates the upward movement of the cell suspension along the inner wall 306. In this regard, the internal cavity 302 may have a substantially conical shape, as illustrated by way of example in FIG. Specifically, the central body region 308 of the separation cup 300 may have a substantially frustoconical shape in this example, and the bottom region 312 of the separation cup 300 is substantially hemispherical in this example. It can have the shape of Accordingly, the upper end 314 of the central body region 308 can be wider than the lower end 316 of the central body region 308 and the diameter D of the inner cavity 302 is downward through the central body region 308 and the lower end 316 of the central body region 308. It can be tapered towards. That is, the diameter D decreases in the direction toward the lower end 316 in the central body region 308.

  Separator cup 300 also includes a shoulder 318 circumscribing upper end 314 of central body region 308 in this example. As seen in FIG. 5, the shoulder 318 defines a neck region 310 above the central body region 308 and a shoulder trap 320 below the neck region 310 in this example. The shoulder trap 320 also circumscribes the upper end 314 of the central body region 308 of the separation cup 300. Accordingly, each of shoulder 318 and shoulder trap 320 can be described as having an annular shape. As can be seen in FIG. 5, the shoulder trap 320 is wider than the neck region 310. In this manner, the shoulder trap 320 of the separation cup 300 collects (ie, captures, localizes, etc.) a relatively high density of material as the separation cup 300 is spun around the central axis. As the separation cup 300 spins about the central axis, the cell suspension moves upward along the inner wall 306 toward the shoulder trap 320. Shoulder trap 320 collects a relatively high density material in the cell suspension, while a relatively low density material in the cell suspension moves upward through the neck region 310. And continue to drain from the separation cup 300 through the opening 322 at the top of the neck region 310. Separation cup 300 may also include an opening 324 circumscribing neck region 310 above neck region 310. Opening portion 324 may have an outwardly extending shape, which is also an upward direction along the inner wall 306 of a relatively low density of material as the separation cup 300 rotates about the central axis. , Facilitating movement out of its top.

  The separation cup 300 also includes a rim 326 that surrounds the opening 324 of the cup 300 and a base 328 at the bottom of the cup 300 in this example. As seen in FIG. 5, an annular notch 330 may be formed on the underside of the rim 326 so that the notch 330 separates the outer wall 332 of the separation cup body 304 from the surface 334 facing inward of the rim wall 336. To do. The base 328 of the separation cup 300 may include one or more coupling members that couple the separation cup 300 to the rotary actuator, and thus, when the separation cup 300 is driven by the rotary actuator. Separation cup 300 is configured to rotate about its central axis. In the illustrated example, the base 328 includes a plurality of legs 337 that are used to secure the separation cup 300 within the installation assembly (220 in FIG. 4A), as further discussed below. Can be done.

  With particular reference to FIG. 6, as described above, the internal cavity 302 comprises a neck region 310, a shoulder trap 320, a central body region 308, and a bottom region 312 in this example from top to bottom. In this example, the neck region 310 can be described as having a truncated cone shape, the shoulder trap 320 can be described as having an annular shape, and the central body region 308 also has a truncated cone shape. The bottom region 312 can then be described as having a hemispherical shape. Thus, from top to bottom, the inner diameter D of the inner cavity 302 can decrease through the neck region 310, increase through the shoulder trap 320, and in this example, the central body region 308 and the bottom region 312. You can decrease through. In this manner, the neck region 310, the central body region 308, and the bottom region 312 can be described as tapered in the downward direction in this example. Accordingly, the shoulder trap 320 can be described as extending laterally toward the outer wall 332 of the cup body 304 in this example. As can be seen in FIG. 6, the contour of the inner wall 306 can follow a somewhat S-shaped curve through the shoulder trap 320.

  When the separation cup 300 is spun around the central axis, centrifugal force acts on the cell suspension retained in the cup 300. As the separation cup 300 spins, the centrifugal force pulls the cell suspension toward the tapered inner wall 306. Due to the conical shape of the inner cavity 302, this cell suspension moves up the inner wall 306 toward the annular shoulder trap 320. When the cell suspension reaches the shoulder trap 320, the relatively high density material in the cell suspension collects in the shoulder trap 320. The relatively low density material in the cell suspension continues to move upwards across the shoulder trap 320, along the neck region 310, and along the outwardly extending opening 324 to separate. Exit the cup 300.

  As can also be seen in FIG. 6, the rim 326 of the separation cup 300 is an annular rim 326 in this example and is folded back to bring the annular notch 330 into a surface 334 facing the inside of the rim wall 336 of the rim 326, It is formed between the outer wall 332 of the main body 304 of the separation cup 300. As shown by way of example in FIG. 6, the rim wall 336 of the rim 326 is substantially parallel to the outer wall 332 of the separation cup body 304, and the notch 330 is located between the rim wall 336 and the outer wall 332.

  Separation cup 300 may have a height of, for example, about 100 millimeters (mm) and an outer diameter of, for example, about 70 mm. The inner cavity 302 can have a depth of, for example, about 70 mm, and the inner diameter D of the conical inner cavity 302 can be, for example, from about 35 mm near the upper end 314 of the central body region 308 to the lower end of the central body region 308. It can be tapered to about 10 mm near 316.

  Referring to FIG. 7, a top perspective view of an example of one embodiment of a separation cup housing 400 is shown. Separator cup housing 400 may include a plurality of installation flanges 402 around base 404 that are used to install isolation cup housing 404 on an installation frame (202 in FIG. 4A). As discussed above, the separation cup (204 in FIG. 4A) is located within the separation cup housing 400. Accordingly, the cup housing 400 includes a central duct 406 defined in this example by a vertical duct wall 408 that continues to the lower interior chamber 410. The duct wall 408 has a cylindrical shape in this example.

  Separation cup 204 can be inserted through central duct 406 so that the lower portion of separation cup 204 (eg, base 328 of FIG. 6) is located within lower interior chamber 410 of cup housing 400. The underside of the separation cup housing 400 is open in this example so that the cup mounting assembly (220 in FIG. 4A) can be received in the lower inner chamber 410 and located in the lower inner chamber 410. The cup housing 400 also includes an annular collection chamber 412 that surrounds the central duct 406 and the separation cup 204.

  The collection chamber 412 receives relatively low density material exiting the separation cup 204 by centrifugal force as the cup 204 spins about the central axis in the central duct 406. The floor 414 of the collection chamber 412 is inclined toward the upper drainage opening 416 formed in the floor 414 of the collection chamber 412. The drainage pipe 418 connects the upper drainage opening 416 to the lower lower drainage opening 420 in the base 404 of the cup housing 400. The lower drainage opening 420 may be disposed over the drainage opening (240 in FIG. 4A) of the installation frame 202. This drain opening receives the drain nozzle (238 in FIG. 4A) as described above. In this manner, the cup housing 400 facilitates removal of low density material exiting the separation cup 204 by centrifugal force.

  FIG. 8 is a side sectional view of the cup housing 400 of FIG. As seen in FIG. 8, the cup housing 400 includes an annular collection chamber 412 formed between the duct wall 408 of the central duct 406 and the outer wall 422 of the cup housing 400. The central duct 406 continues to a lower internal chamber 410 that houses the cup mounting assembly 220, as discussed further below. The floor 414 of the collection chamber 412 is inclined toward the upper drain opening 416 and the drain pipe 418.

  Referring now to FIG. 9, a side cross-sectional view of the upper portion of the apparatus 200 for separating different density materials of FIG. 4A is shown in an assembled configuration. As can be seen in FIG. 9, the separation cup 300 is located in the central duct 406 of the cup housing 400. The duct wall 408 of the central duct 406 separates the separation cup 300 from the collection chamber 412 of the cup housing 400. The separation cup 300 passes through the central duct 406 of the cup housing 400 so that the lower portion of the separation cup 300 is located in the lower inner chamber 410 of the cup housing 400 along with the cup mounting assembly 220. The lower inner chamber 410 of the cup housing 400 is discussed in further detail below with reference to FIG.

  Further, the top of the duct wall 408 is received in the notch 330 of the rim 326 that surrounds the top of the separation cup 300 so that the rim wall 334 of the rim 326 and the duct wall 408 of the central duct 406 are illustrated by way of example. As shown in FIG. In this manner, the notch 330 prevents fluid discharged from the separation cup 300 from contacting the outer wall 332 of the separation cup 300. Further, the housing cap 242 may cover the top of the cup housing 400 for the purpose of confining the separation cup 300 located within the housing 400. The pipetter opening 244 of the housing cap 242 is located above the opening 208 of the separation cup 204. In this manner, a pipettor can be inserted through the opening 244 in the housing cap 242 for the purpose of sucking the medium from the separation cup 300 and dispensing the medium into the separation cup 300. As seen in FIG. 9, this configuration allows the pipettor to aspirate and dispense media even when the separation cup 300 is spinning.

  The configuration of the separation cup 300 and cup housing 400 shown by way of example in FIG. 9 facilitates the separation of relatively high density materials and relatively low density materials using centrifugal forces. As explained above, when the cup 300 is in the cup housing 400, the separation cup 300 can be spun around the central axis. The relatively high density material is trapped in the shoulder trap 320 of the inner cavity 302 and the relatively low density material exits through the outwardly extending opening 324 of the cup 300. Accordingly, the annular collection chamber 412 of the cup housing 400 that surrounds the separation cup 300 receives a relatively low density of material exiting the separation cup 300 by centrifugal force.

  Referring now to FIG. 10, a perspective view of an example of one embodiment of a cup installation assembly 500 is shown. The cup mounting assembly 500 includes a rotor mount 502, a biasing spring 504, a support pad 506, and a cup mount 508 in this example. In this example, the biasing spring 504 and support pad 506 are received within the rotor mount 502 and the cup mount 508 is secured to the rotor mount 502 (eg, by screws or other attachment means). In FIG. 10, the upper surface of the rotor mount 502 is shown, and the lower surfaces of the support pad 506 and the cup mount 508 are shown.

  The rotor mount 502 includes, in this example, a recess 510 that receives the biasing spring 504 and the support pad 506, and a central duct 512 for reception of the shaft 514 of the support pad 506. In this example, the biasing spring 504 is disposed so as to surround the wall 516 of the central duct 512 of the rotor mount 502. The biasing spring 504 is also arranged to surround the shaft 514 of the support pad 506 in this example. The support pad 506 also includes a recess 518 formed on the lower surface thereof for receiving the biasing spring 504. In this configuration, the biasing spring 504 biases the support pad 506 in a direction away from the rotor mount 502. When the biasing spring 504 is loaded, the central duct 512 of the rotor mount 502 receives the shaft 514 of the support pad 506. The rotor mount 502 is coupled to the rotor assembly (222 in FIG. 4A) and receives the rotational force provided by the rotary actuator (206 in FIG. 4A).

  Separator cup mount 508 secures separator cup 300 in this example when separator cup (300 in FIG. 5) is placed in the cup housing (400 in FIG. 7). As can be seen in FIG. 10, the separation cup mount 508 comprises a set of internal flanges 520 that are formed through the cup mount 508 for the base of the separation cup 300 (328 in FIG. 5). An opening 522 is defined. The base opening 522 defined by the inner flange 520 has a shape that matches the base 328 of the separation cup 300. Accordingly, the base opening 522 of the cup mount 508 includes a respective opening 524 for receiving the foot of the base 328 of the separation cup 300 (337 in FIG. 5). The cup mount 508 also includes a recess 526 for the base 328 of the separation cup 300 formed in the lower surface of the inner flange 520 in this example. The shape of the base recess 526 on the lower surface of the inner flange 520 also matches the shape of the base 328 of the separation cup 300 in this example. Accordingly, each inner flange 520 is each provided with a recess 528 for receiving one of the legs 337 of the base 328 of the separation cup 300 in this example.

  However, the shape of the base recess 526 on the lower surface of the cup mount 508 rotates with respect to the shape of the base opening 522 of the cup mount 508. Accordingly, the base 328 of the separation cup 300 can be inserted through the opening 522 of the cup mount 508, and the legs 337 of the base 328 of the separation cup 300 are aligned with the foot recesses 528 formed in the lower surface of the inner flange 520. Can be rotated so that it is received in the foot recess 528. Accordingly, the recess 528 formed in the lower surface of the inner flange 520 can fix and lock the foot 337 to the base 328 of the separation cup 300. In this manner, the cup mount 508 can apply a rotational force provided by the rotary actuator 206 to the separation cup 300 for the purpose of spinning the cup 300 about the central axis.

  The rotor mount 502 may also include a respective recess 530 for receiving the foot 337 in the base 328 of the separation cup 300. As can be seen in FIG. 10, the foot recess 530 is clockwise in the rotor mount 502 for the purpose of aligning the foot 337 of the base 328 of the separation cup 300 with the foot opening 524 or foot recess 528 of the cup mount 508, respectively. And is sized and shaped to allow it to rotate counterclockwise. In this manner, the separating cup 300 is unidirectional to secure the cup 300 to the cup mount 508 by aligning the foot 337 of the base 328 of the cup 300 with the foot recess 528 formed in the lower surface of the inner flange 520. Can be rotated. Similarly, the separation cup 300 can be rotated in the reverse direction to remove the cup 300 from the cup mount 508 by aligning the foot 337 of the base 328 of the cup 300 with the foot opening 524 defined by the internal flange 520. obtain. The rotary actuator (206 in FIG. 4A) may include a brake to prevent rotation of the rotary actuator when the separation cup 300 is secured or removed within the cup installation assembly 500.

  Referring to FIG. 11, a side cross-sectional view of the lower portion of the apparatus 200 for separating different density materials of FIG. 4A is shown in an assembled configuration. As seen in FIG. 11, the lower part of the separation cup 300 and the cup installation assembly 500 are located in the lower inner chamber 410 of the separation cup housing 400.

  When the separation cup 300 is inserted into the separation housing 400, the cup 300 passes through the central duct 406 until the base 328 of the cup 300 engages the cup mounting assembly 500. Separation cup 300 can be rotated until foot 337 of base 328 of cup 300 is aligned with the foot opening (524 in FIG. 10) of cup mount 508. With the foot 337 of the base 328 of the separation cup 300 aligned with the foot opening 524 of the cup mount 508, the cup 300 can be pushed downward through the opening (522 in FIG. 10) of the cup mount 508. When the separation cup 300 is pushed through the opening 522 of the cup mount 508, the base 328 of the cup 300 engages the support pad 506 and loads the biasing spring 504. The foot 337 of the base 328 of the separation cup 300 is also received in the foot recess 530 of the rotor mount 502 when the cup 300 is pushed through the opening 522 of the cup mount 508.

  Once the foot 337 of the base 328 of the separation cup 300 passes through the opening 522 of the cup mount 508, the foot of the base 328 of the cup 300 is formed on the lower surface of the inner flange 520 of the cup mount 508. It can be rotated within the foot recess 530 of the rotor mount 502 to align with the recess 528. The separation cup 300 can then be released so that the biasing spring 504 is unloaded and pushes the separation cup 300 upward and a recess formed in the lower surface of the inner flange 520 of the cup mount 508 (see FIG. 10 526). A foot recess 528 formed in the lower surface of the inner flange 520 receives the foot 337 of the base 328 of the separation cup 300. The biasing spring 504 continues to bias the support pad 506 against the base 328 of the separation cup 300, thereby fixing the cup 300 in a recess 526 formed on the lower surface of the inner flange 520 of the cup mount 508. . Fixed in the recess 526, the cup mount 508 can apply the rotational force provided by the rotary actuator (206 in FIG. 4A) to the separation cup 300 as described above.

  The separation cup 300 reverses this procedure (ie, pushes the separation cup 300 downward so that the foot recess 530 of the rotor mount 502 receives the foot 337 of the base 328 of the cup 300; By rotating the cup 300 until the foot 337 is aligned with the foot opening 524 of the cup mount 508; and lifting the cup 300 from the cup mounting assembly 500 through the opening 522 of the cup mount 508). obtain.

  Referring now to FIG. 12, an example of an alternative embodiment of a separation cup 600 is shown. This alternative embodiment of the separation cup 600 includes one or more passages 602 formed through the body 304 of the cup 600, from the lower end 316 of the central body region 308, near the annular shoulder trap 320. The flow path to the upper end 314 of the central body region 308 is defined. Each of the passages 602 includes a longitudinal bore 604 formed through the body 304 of the separation cup 600, which longitudinal bore 604 is formed in the inner wall 306 near the lower end 316 of the central body region 308. Is connected to the upper aperture 608 formed on the inner wall 306 near the upper end 314 of the central body region 308 and below the shoulder trap 320.

  As the separation cup 600 spins, centrifugal force drives the cell suspension toward the lower aperture 606 formed in the inner wall 306 near the lower end 316 of the central body region 308. This centrifugal force drives the cell suspension through the lower aperture 606 and enters the bore 604 in the passage 602. The centrifugal force then drives the cell suspension upward through the bore 604 in the passage 602 and out of the upper aperture 608 below the shoulder trap 320. As described above, when this cell suspension exits the upper aperture 608, relatively high density material accumulates in the shoulder trap 320, while relatively low density material passes through the shoulder trap 320. Continue to go out of the cup 600.

  As described above, the cell suspension may include cells (relatively dense material) suspended in a medium, such as a buffer solution (relatively dense material). The passage 602 of the separation cup 600 is advantageously in this alternative example as these cells move in the buffer solution as they move upwardly through the passage 602 toward the shoulder trap 320. To help ensure that it remains immersed.

  An alternative embodiment of the separator cup 600, shown by way of example in FIG. 12, comprises six passages 602 from the lower end 316 to the upper end 314 of the central body region 308. Each of the passages 602 includes a longitudinal bore 604 that in this example connects a lower aperture 606 formed in the inner wall 306 to an upper aperture 608 formed in the inner wall 306. Also as seen in FIG. 12, the passage 602 can be angled with respect to the body 304 of the separation cup 600 in this example.

  Further, the passageway 602 can include various regions that define respective flow paths. As shown by way of example in FIG. 12, the passage 602 may comprise a lower elbow region 610 connected to the lower aperture 606; the lower elbow region 610 continues to the inclined middle region 612; The upper elbow region 614 continues to the vertical top region 616; and the vertical top region 616 is connected to the upper aperture 608.

  The upper aperture 608 and the lower aperture 606 may have a circular or elliptical shape, for example. Accordingly, the cross-section of the bore 604 of the passage 602 can also have a circular or elliptical shape. The width of the apertures 606 and 608 can be, for example, about 1 mm, and the width of the bore 604 in the passage 602 can also be, for example, about 1 mm. The passage 602 may have a length of about 40 mm, for example.

  A method of separating different density materials using the separation cup has also been developed. This method can be used to prepare cells for use in biological assays such as those performed by automated high-throughput electrophysiological measurement systems. As will be apparent from the present disclosure, the method may be automated and therefore implemented using a suitable automation system (eg, the system 100 described above and illustrated by way of example in FIGS. 1 and 2). Can be done.

  Referring to FIG. 13, a flowchart 700 of example method steps for separating materials of different densities is shown. In this example, the method has been described in terms of preparing cells for biological assays. However, it should be understood that this method may be used in other respects to separate other types of different density materials.

  Initially, a pipettor can aspirate a cell solution from the cell holding station and dispense the cell solution into a separation cup (step 702). For example, a solution of about 15 milliliters (ml) to about 30 ml of cells can be dispensed into the separation cup. The pipetter can dispense the solution of cells into the separation cup, for example, at a rate of about 100 microliters (μl) per second (s).

  The separation cup can then be rotated about the central axis for the purpose of removing relatively low density material from the cup as described above (step 704). The separation cup can be rotated at a spin rate of about 3000 revolutions per minute (rpm) in this example for the purpose of removing relatively low density material.

  After the relatively low density material has been removed from the separation cup, the pipetter can dispense the desired solution (eg, wash buffer) into the cup (step 706). For example, about 20 ml to about 30 ml of the desired solution can be dispensed into a separation cup. As above, the pipettor can dispense the desired solution into the separation cup at a rate of about 100 μl / s. The pipetter can deliver the desired solution to the separation cup while the cup is still spinning.

  Once the desired solution has been delivered to the separation cup, the cup can be agitated to remove the relatively dense material from the shoulder trap and place the relatively dense material in the desired solution (step 708). . In this manner, the method ensures that substantially all of the relatively dense material enters the desired solution.

  Agitation of the separation cup to mix the desired solution and cells can include controlling the duration, direction, speed, acceleration, and deceleration of rotation. As described above, when the rotary actuator drives the rotation of the separation cup, the control module may control the rotary actuator. Referring to FIG. 14, a flowchart 800 of example method steps for automatically stirring a separation cup is shown. It will be appreciated that any combination or subset of the agitation steps discussed below can be selectively performed to agitate the separation cup. Initially, the separation cup can be accelerated forward (eg, clockwise) at a predetermined forward acceleration rate (step 802), and the predetermined forward spin speed can be maintained for a predetermined duration ( Step 804). After maintaining this forward spin rate, the separation cup can be decelerated at a predetermined forward deceleration rate (step 806). The predetermined forward acceleration rate can be, for example, 1000 rpm (rpm / s) per second; the predetermined forward spin speed can be, for example, 3000 rpm; the predetermined duration can be, for example, 100 s And the predetermined forward deceleration rate can be, for example, 1000 rpm / s.

  The separation cup can then be accelerated backwards (eg, counterclockwise) at a predetermined backward acceleration rate (step 808), and the predetermined backward spin rate can be maintained for a predetermined duration. (Step 810). After maintaining this backward spin rate, the separation cup can be decelerated at a predetermined backward deceleration rate (step 812). The predetermined backward acceleration rate can be, for example, 1000 rpm / s; the predetermined backward spin rate can be, for example, 3000 rpm; the predetermined duration can be, for example, 100 s; and the The predetermined backward deceleration rate may be 1000 rpm / s, for example.

  After spinning the separation cup forward and backward as described above, a pipettor can be used to crush or titrate the cell solution (step 814), thereby providing the desired level of homogeneity. Achieve. Trituration or titration of the cell solution may include aspirating the cell solution from the separation cup with a pipetter and dispensing the cell solution back into the separation cup. The pipetter can aspirate and dispense the solution a predetermined number of times (eg, 10 times) when grinding or titrating the solution of cells. If more uniform homogeneity is desired, further grinding or titration steps can be performed.

  Generally, terms such as “contact”, “communicate”, “communicate with ...” and “communicate with ...” (eg, the first component “contacts” the second component) ”,“ Communicate ”,“ contacting ”or“ communicating ”) is a structural relationship, a functional relationship, a mechanical relationship, an electrical relationship between two or more components or elements Used herein to indicate a relationship, signal relationship, optical relationship, magnetic relationship, electromagnetic relationship, ionic relationship, or fluid relationship. Thus, the fact that one component is described as being in communication (communication) with a second component is that additional components can exist between the first component and the second component. It is not intended to exclude the possibility and / or the possibility of operably related or engaged.

  The description of the above embodiments has been presented for purposes of illustration and description. The above description is not exclusive and is not intended to limit the invention to the precise form disclosed. Modifications and changes are possible in light of the above description or may be obtained from practice of the invention. The claims and their equivalents define the scope of the invention.

204, 300, 600 Device 304 Cup body 302 Internal cavity 306 Inner wall 308 Central body region 318 Internal shoulder 310 Neck region 320 Shoulder trap 322 Opening 200 Device

Claims (15)

A device (300) for separating materials of different density, the device comprising:
A cup body (304) having an internal cavity (302) configured to hold a medium;
An inner wall (306) defining a central body region (308) having an upper end and a lower end, the upper end being wider than the lower end, an inner wall (306); and circumscribing the upper end of the central body region (308) An internal shoulder (318), wherein the internal shoulder defines a neck region (310) above the central body region and a shoulder trap (320) below the neck region, the shoulder trap An inner shoulder (318) circumscribing the upper end of the central body region
Comprising:
The shoulder trap (320) is wider than the neck region (310) so that when the device is spun around a central axis, the media is directed upward along the inner wall (306) and the shoulder Moving toward the trap (320), a relatively high density material in the medium is collected in the shoulder trap, and a relatively low density material in the medium is an opening above the neck region. Device (300) ejected from the device through section (322).   The inner cavity (302) in the central body region (308) has a substantially conical shape, so that the diameter of the inner cavity (302) in the central body region is the same as that of the central body region. The device of claim 1, wherein the device tapers downward toward the lower end.   The coupling member (328) configured to couple the device to a rotary actuator (206), the cup body (304) being rotatable about a central axis of the cup body. 2. The device according to 2.   The device of claim 1, 2 or 3, wherein the inner wall (306) defines a bottom region below the central body region (308), and the bottom region has a substantially hemispherical shape.   Above the neck region (310) is further provided an opening (324) defining the opening (322), through which the relatively low density material is discharged, the opening being 5. A device according to any one of the preceding claims, circumscribing the neck region and extending outward in a direction away from the neck region.   And further comprising one or more passages (602) formed through the cup body (304) from the lower end of the central body region (308) to the upper end of the central body region, each of the one or more passages , Defining a flow path from the lower end to the upper end of the central body region, and in particular, each of the one or more passageways (602) is a longitudinal bore (604) formed through the cup body (304). The longitudinal bore includes a lower aperture formed in the inner wall (306) near the lower end of the central body region (308) and a shoulder trap near the upper end of the central body region. 6. A device according to any one of the preceding claims, connected to an upper aperture formed in the inner wall below (320). A method of separating a cell suspension from a starting mixture, the starting mixture comprising a relatively low density buffer solution and a relatively high density of cells, the method comprising:
Dispensing the starting mixture to a device (204, 300, 600), the device comprising a cup body (304) having an internal cavity (302) for holding the starting mixture; A device, in particular a device according to any one of claims 1 to 6;
Spinning the device about a central axis so that the starting mixture is directed upward along the inner wall (306) of the cup body (304) and circumscribing the inner cavity (320) ) So that the relatively high density cells in the starting mixture are collected in the shoulder trap and at least some of the relatively low density buffer solution in the starting mixture is Evacuating from the device through an opening (322) in the cup body;
Dispensing a desired solution into the internal cavity (302) of the cup body (304) after draining at least some of the relatively low density buffer solution; and the desired solution and the relatively high Stirring the device such that the cells of density are mixed together. Stirring the device (300) includes:
Accelerating the device forward at a predetermined forward acceleration rate;
Maintaining a predetermined forward spin rate for a first predetermined amount of time;
Decelerating the device at a predetermined forward deceleration rate;
Accelerating the device backwards at a predetermined backward acceleration rate;
Maintaining a predetermined backward spin rate for a second predetermined amount of time;
8. The method of claim 7, comprising: decelerating the device at a predetermined backward deceleration rate; and at least one of grinding or titrating the desired solution and the relatively dense cells with a pipettor. the method of. An apparatus (200) for separating materials of different densities, the apparatus comprising:
Separation cup (204, 300), in particular a separation device according to any one of the preceding claims, wherein the separation cup is configured to hold an internal cavity (302). And a shoulder trap (320) circumscribing the internal cavity, the shoulder trap being a relatively dense material in the medium when the separation cup is spun around a central axis Is collected in the shoulder trap (320) and a relatively low density material in the medium is configured to be discharged from the separation cup (204, 300). And a rotation actuator (206) coupled to the separation cup, the rotation actuator being configured to spin the separation cup around a central axis of the separation cup Actuator (206)
An apparatus (200) comprising: The separation cup (204, 300) is:
An inner wall (306) defining a central body region (308) having an upper end and a lower end, the upper end being wider than the lower end, an inner wall (306); and circumscribing the upper end of the central body region (308) An inner shoulder (318), the inner shoulder further comprising an inner shoulder defining the shoulder trap (320) circumscribing the inner cavity (302) and a neck region above the central body region;
In particular, the internal cavity (302) in the central body region (308) of the separation cup has a substantially conical shape so that the diameter of the internal cavity in the central body region is equal to the central body. The apparatus of claim 9, wherein the apparatus tapers downward toward the lower end of the region. The diameter of the internal cavity (302) of the separation cup (204, 300) increases downward through the shoulder trap (320), and the diameter of the internal cavity of the separation cup is determined by the neck And / or the separation cup (204, 300) further comprises an opening (324) circumscribing the neck region above the neck region (310). And the opening (324) extends outwardly away from the neck region,
The apparatus according to claim 9 or 10. The separation cup (204, 300, 600) further includes one or more passages (602) formed through the separation cup from the lower end of the central body region (308) to the upper end of the central body region. Each of the one or more passageways (602) defines a flow path from the lower end to the upper end of the central body region of the separation cup;
12. A device according to claim 9, 10 or 11.   Each of the one or more passageways (602) of the separation cup (204, 300, 600) comprises a longitudinal bore (604), the longitudinal bore being in the central body region (308). A lower aperture (606) formed in the inner wall (306) near the lower end and an upper aperture (608) formed in the inner wall near the upper end of the central body region and below the shoulder trap (320). The device of claim 12, connected to the device.   A housing (400) surrounding the separation cup (204, 300, 600), the housing comprising a collection chamber (412), circumscribing the opening (324) of the separation cup; As a result, when the separation cup is spun around the central axis, the collection chamber receives the relatively low density material discharged from the separation cup, and in particular the collection chamber (412) Floor (414) is inclined towards the drain (416, 418) so that the relatively low density material discharged from the separation cup can be drained from the collection chamber. 14. The device according to any one of claims 9 to 13, wherein:   An installation assembly (220, 500) that couples the separation cup (204, 300, 600) to the rotary actuator (206), the separation cup being removably fixable within the installation assembly. Item 15. The apparatus according to any one of Items 9 to 14.