Patent Publication Number: US-11027284-B2

Title: Well plate mixing apparatus

Description:
CROSS-REFERENCE TO RELATED APPLICATION 
     This application claims the benefit of U.S. provisional patent application No. 62/611,005, filed Dec. 28, 2017. The content of this application is incorporated by reference in its entirety. 
    
    
     FIELD OF THE INVENTION 
     Aspects of this invention relate generally to a mixing apparatus for a well plate, and more particularly, to a mixing apparatus that uses an electromagnet assembly to move a well plate with respect to a fixed sensor mount. 
     BACKGROUND OF THE INVENTION 
     Well plates have been and are the industry standard for many types of chemical and biological testing and screenings. Prior art well mixing devices include vibrational plates and orbital motion plates. Changes in the configuration of well plates has decreased the effectiveness of such methods. The density of well plates has transitioned from 48 to 96 to 386 to 1536 wells, drastically decreasing the volume to surface area ratio of each well. As microplate well volumes decrease, variables such as surface tension and the aspect ratio of taller, thinner wells have decreased the effectiveness of traditional mixing techniques. As the diameter of the well decreases, the Reynolds number decreases, resulting in viscous forces becoming more dominant than convective forces in the well, reducing the effectiveness of efforts to mix the contents of the well. Therefore, movement of the well plate is not a sufficient method alone to mix the contents of the well. Pipetting the solution into and out of the well multiple times is effective in mixing the contents of the well, however it is not automated and is not reproducible. Ultrasonic mixing has also been used, but it can introduce heat into the well, which can cause damage to certain molecular species like proteins, DNA, and cells, which are of key interest to many of the well plate studies. 
     It would be desirable to provide a well mixing apparatus that reduces or overcomes some or all of the difficulties inherent in prior known processes. Particular objects and advantages will be apparent to those skilled in the art, that is, those who are knowledgeable or experienced in this field of technology, in view of the following disclosure and detailed description of certain embodiments. 
     SUMMARY 
     In accordance with a first aspect, a mixing apparatus includes a well plate assembly including a fixed support, and a well movable with respect to the fixed support. A fixed sensor mount has a first portion disposed above the well and a second portion disposed within the well. A plurality of electromagnets are operable to move the well plate assembly vertically with respect to the fixed sensor mount and the fixed support. 
     In accordance with another aspect, an apparatus for mixing a well plate includes a well plate assembly having a fixed support, a base plate having an upper base plate portion spaced from the fixed support and a lower base plate portion spaced from the fixed support, and a well plate including a plurality of wells and movable with respect to the fixed support. Each of a plurality of fixed sensor mounts has a first portion disposed above a selected one of the plurality of wells and a second portion disposed within the selected well. Each of a plurality of sensors is secured to a selected one of the sensor mounts. An electromagnet assembly is operable to move the well plate assembly vertically with respect to the fixed support and the fixed sensor mounts. The electromagnet assembly includes a plurality of electromagnets, each of which includes a magnet housing, an electromagnet fixed in the magnet housing, and a permanent magnet movable with respect to the magnet housing and secured to the base plate. A spring assembly includes a plurality of base plate springs captured between the base plate and the fixed support. 
     These and additional features and advantages disclosed here will be further understood from the following detailed disclosure of certain embodiments, the drawings thereof, and from the claims. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
       The foregoing and other features and advantages of the present embodiments will be more fully understood from the following detailed description of illustrative embodiments taken in conjunction with the accompanying drawings in which: 
         FIG. 1  is an elevation view of a mixing apparatus for a well plate. 
         FIG. 2  is a section view, partially broken away, of an electromagnet of the mixing apparatus of  FIG. 1 . 
         FIG. 3  is a section view, partially broken away, a spring assembly of the mixing apparatus of  FIG. 1 . 
         FIG. 4  is a schematic front elevation view of a sensor mount in a well of the mixing apparatus of  FIG. 1 . 
         FIG. 5  is a schematic side elevation view of a sensor mount in a well of the mixing apparatus of  FIG. 1 . 
         FIG. 6  is a plan view of a sensor mount in a well of the mixing apparatus of  FIG. 1 . 
         FIG. 7  is a schematic front elevation view of an alternative embodiment of a sensor mount in a well of the mixing apparatus of  FIG. 1 . 
         FIG. 8  is a schematic front elevation view of another alternative embodiment of a sensor mount in a well of the mixing apparatus of  FIG. 1 . 
     
    
    
     The figures referred to above are not drawn necessarily to scale, should be understood to provide a representation of particular embodiments, and are merely conceptual in nature and illustrative of the principles involved. Some features of the mixing apparatus depicted in the drawings have been enlarged or distorted relative to others to facilitate explanation and understanding. The same reference numbers are used in the drawings for similar or identical components and features shown in various alternative embodiments. Mixing apparatuses for well plates as disclosed herein would have configurations and components determined, in part, by the intended application and environment in which they are used. 
     DETAILED DESCRIPTION OF EMBODIMENTS 
       FIG. 1  illustrates a representative mixing apparatus  10  that may be used to mix the contents of a well plate assembly  12 . Well plate assembly  12  includes one or more wells  14  disposed in a well plate  16 . In certain embodiments, well plate  16  may have as many as 1536 and at least 24 wells  14 . It is to be appreciated that well plate  16  may include any number of wells  14 . 
     One or more fixed sensor mounts  18  are positioned such that each sensor mount  18  has a first portion  20  disposed above a well  14  and a second portion  22  is disposed within the well  14 . As discussed in greater detail below, sensor mounts  18  serve to mix the contents of well  14 . In certain embodiments, sensor mounts  18  may be formed of plastic, metal, ceramic, silicon, glass or a PCB. 
     A base plate  24  is positioned below and in abutting relationship with well plate  16 . Well plate  16  and base plate  24  are coupled to one another so that if base plate  24  moves, well plate  16  moves accordingly. They can be coupled by any number of means including gravity acting on well plate  16 , a clamp, a fastener, etc. In some embodiments, mixing apparatus  10  is specifically designed to facilitate quick removal and/or replacement of well plate assembly  12 . 
     An electromagnet assembly  25  includes a plurality of electromagnets  26 , described in greater detail below, are positioned beneath base plate  24 . Electromagnets  26  serve to move well plate assembly  12  with respect to fixed support  30 , thereby moving each well  14  with respect to the fixed sensor mount  18  disposed therein. In certain embodiments, well plate assembly  12  moves primarily vertically, or in the Z direction, (as indicated by arrows A) with respect to fixed support  30 . It is to be appreciated that in certain embodiments, discussed in greater detail below, electromagnets  26  serve to move well plate assembly  12  horizontally as well, in the X and Y directions. 
     A portion  28  of a fixed support  30  that surrounds well plate assembly  12  is received in a recess  32  formed into base plate  24  and defining an upper base plate portion  24   a  and a lower base plate portion  24   b . Portion  28  of fixed support is positioned within recess  30  such that it does not contact well plate assembly  12 . Thus, upper base plate portion  24   a  and lower base plate portion  24   b  are each spaced from fixed support  30 , allowing movement of base plate  24  and well plate assembly  12  with respect to fixed support  30 . 
     In use, when electromagnets  26  are activated, well plate assembly  12  moves with respect to both fixed support  30  and sensor mounts  18 . As wells  14  move with respect to sensor mounts  18 , the liquid within wells  14  is mixed not only by the movement of well  14 , but also by the movement of sensor mounts  18  within wells  14 , as described in greater detail below. 
       FIG. 2  illustrates an electromagnet  26  in greater detail. Electromagnet  26  includes an electromagnetic coil  40  disposed within a magnet housing  42 . A permanent magnet  44  is positioned partially within magnet housing  42  and partially within a magnet recess  45  formed in a bottom surface of base plate  24 . Permanent magnet  44  is secured to base plate  24  such that when permanent magnet  44  moves, it causes well plate  16  and, naturally, well  14  to move as well. 
     In certain embodiments, electromagnet assembly  25  includes four electromagnets  26  positioned directly beneath base plate  24  and beneath well plate assembly  12 , with only two being visible in  FIG. 1 . It is to be appreciated that electromagnet assembly  25  can include any desired number of electromagnets  26 . 
     Another portion of electromagnet assembly  25  is shown in  FIG. 3 , where it can be seen that spring assembly  46  is housed within base plate  24  underneath well plate assembly  12 , specifically between upper base plate portion  24   a  and lower base plate portion  24   b . Spring assembly  46  works with electromagnets  26  to move base plate  24 , well plate  16 , and wells  14  with respect to fixed support  30 . Well plate  16  and wells  14  move primarily vertically, or in the Z direction, (as indicated by arrows C) with respect to fixed support  30  with and against the biasing action of spring assembly  46 . As noted above, well plate  16  and wells  14  of well plate assembly  12  can move horizontally as well, in the X and Y directions. 
     Spring assembly  46  includes a first spring  48  that is captured between lower base plate portion  24   b  and fixed support  30 . A first lower end  50  of first spring  48  is seated in a base plate recess  52  formed in an upper surface of lower base plate portion  24   b . A second upper end  54  of first spring  48  is seated in a first fixed support recess  56  formed in a bottom surface of fixed support  30 . 
     A second spring  58  is captured between upper base plate portion  24   a  and fixed support  30 . A first lower end  60  of second spring  58  is seated in a second fixed support recess  62  formed in an upper surface of fixed support  30 . A second upper end  64  of second spring  58  is seated in a well plate recess  56  formed in a bottom surface of upper base plate portion  24   a.    
     First spring  48  is configured to keep base plate  24  spaced apart a first distance D from fixed support  30  when well plate assembly  12  is in an at rest condition. Similarly, second spring  58  is configured to keep well plate  16  spaced apart from fixed support  30  a second distance E when well plate assembly  12  is in an at rest condition. Distances D and E are the maximum distances that well plate  16  and upper and lower base plate portions  24   a ,  24   b  can move up and down, respectively, in the vertical, or Z, direction. In certain embodiments, distances D and E are each approximately 0.5 mm, providing for a total vertical travel distance for well plate assembly  12  and, therefore, wells  14 , of approximately 1 mm. 
     The term “approximately” as used herein is meant to mean close to, or about a particular value, within the constraints of sensible, commercial engineering objectives, costs, manufacturing tolerances, and capabilities in the field of mixing apparatus manufacturing and use. Similarly, the term “substantially” as used herein is meant to mean mostly, or almost the same as, within the constraints of sensible, commercial engineering objectives, costs, manufacturing tolerances, and capabilities. 
     When electromagnet  26  is activated, electromagnetic coil  40  is alternately energized and de-energized, alternately moving permanent magnet  44 , and therefore, well plate assembly  12 , toward and away from electromagnetic coil  40  and magnet housing  42  against the biasing actions of first spring  48  and second spring  58  in the direction of arrows A seen in  FIG. 1  and arrows B seen in  FIG. 2 . 
     This cycling or oscillating of electromagnet  26  causes the vertical movement of well plate assembly  12  and, therefore wells  14 , which can be seen more clearly in  FIGS. 4-6 . Wells  14  include liquid with contents to be sensed, as denoted by liquid level line  66 . As well  14  moves vertically in the direction of arrows A, sensor mount  18  is stationary causing relative movement compared to well  14 , such that second portion  22  of sensor mount  18 , which is disposed within the liquid, mixes the contents of well  14 . In certain embodiments, electromagnet  26  may oscillate at a rate of approximately 30 Hz. 
     In certain embodiments, the plurality of electromagnets  26  all operate simultaneously in phase so that well plate assembly  12  and wells  14  move only vertically in the Z direction. In other embodiments, electromagnets  26  may operate out of phase with one another. In such an embodiment, well plate assembly  12  and wells  14  do not move only vertically. Rather, in such an embodiment, well plate assembly  12  and wells  14  move with a rocking or orbital motion, providing movement horizontally in the X and Y directions (seen in  FIG. 6 ) as well as vertically in the Z direction. It is to be appreciated, however, that in such an embodiment, well plate assembly  12  and wells  14  move primarily vertically in the Z direction. In other words, in such an embodiment, well plate assembly  12  and wells  14  move a larger distance vertically in the Z direction than they move horizontally in the X and Y directions. 
     The relative movement of well  14  with respect to stationary sensor mount  18  creates two types of mixing of the fluid in well  14 . The first type of mixing is shear mixing that occurs along the surface of sensor mount  18 . This shear mixing is caused by the no-slip boundary condition that occurs at the interface of the liquid in well  14  and sensor mount  18 . The liquid next to sensor mount  18  moves at the same velocity as the stationary sensor mount, while the liquid in the rest of well  14  moves at the same velocity as well  14 , creating a large velocity gradient in the liquid of well  14 . The large velocity gradient in the liquid is the best condition for mixing. Since the large velocity gradient and, therefore, the best mixing conditions are next to the surface of sensor mount  18 , this mixing is especially useful for the addition of certain molecular species from sensor mount  18  to the contents of well  14 , or for movement of a molecular species from well  14  to the surface of sensor mount  18 . In a preferred embodiment, sensor mount  18  may be used for sensing of the concentration of the molecular species next to sensor mount  18 . In this case, the purpose of mixing is so that the rate any change in the species concentration at the surface is limited by reaction kinetics, not diffusion kinetics. 
     The second kind of mixing is due to the liquid displaced by the volume of sensor mount  18 , which will cause large scale displacement currents and eddies in well  14 , greatly increasing the rate of mixing. These displacement currents are useful for mixing the entirety of well  14 . Both of these types of mixing will equilibrate the concentration of molecular species throughout the entirety of well  14 . 
     The combination of positioning sensor mount  18  in well  14  and the mostly vertical motion of well  14  relative to sensor mount  18  provides good, thorough, and efficient mixing. Without sensor mount  18 , or another nonmoving object in well  14 , there will be no shear mixing and no displacement mixing within well  14 . Merely creating vertical movement of well  14  through the operation of electromagnets  16  will not thoroughly mix the contents of well  14 . At the same time, without the predominantly vertical motion of well  14 , there will also not be thorough mixing of the contents of well  14  since there will be no velocity gradient near the surface of sensor mount  18 . In such a case, only diffusion could equilibrate the molecular species concentration from near the surface of sensor mount  18  to the rest of well  14 . 
     The use of sensor mount  18  in conjunction with electromagnet assembly  25  to mix the contents of well  14  provides an easy to use and highly repeatable way of thoroughly mixing the contents of well  14 . Further, such a mixing apparatus imparts low levels of energy to the contents of well  14 , thereby decreasing the chance of damaging the contents of well  14 . 
     It has been discovered that the use of sensor mount  18  in conjunction with electromagnet assembly  25  to mix the contents of well  14  is much more effective than the mixing of well  14  would be when using electromagnet assembly  25  alone. In one experiment it was found that thorough mixing to reach equilibrium in well  14  using just electromagnet assembly  25  (which provides only movement of well  14 ) took approximately one hour, while equilibrium was reached in well  14  using both sensor mount  18  and electromagnet assembly  25  (which provides movement of well  14 , shear mixing, and displacement) within approximately 3 seconds. This provides a greater than 1000× improvement using both sensor mount and electromagnet assembly  25  as compared to using electromagnet assembly  25  alone. 
     In the embodiment illustrated in  FIGS. 4-6 , sensor mount  18  has the shape of an elongate plate. Thus, in such an embodiment sensor mount  18  has a length L, a width W, and a depth D, where the length L is significantly larger than the width W, and the width W is significantly larger than the depth D. This produces a relatively thin and long plate, providing a large ratio of surface area to volume for sensor mount  18 , which enhances the mixing capability of sensor mount  18  within well  14 . 
     In certain embodiments, sensor mount  18  has a length L of between approximately 2 mm and approximately 10 mm a width W of between approximately 1 mm and approximately 15 mm, and a depth of between approximately 0.2 mm and approximately 3.2 mm. Well  14  may have a diameter of between approximately 1.7 mm and approximately 15.6 mm, and a height of between approximately 4.8 mm and approximately 11.0 mm. 
     In another embodiment for a standard 96 well plate, sensor mount  18  has a length L of between approximately 6 mm and approximately 12 mm, a width W of between approximately 4.8 mm and approximately 5.2 mm, and a depth of between approximately 1.4 mm and approximately 1.8 mm. Well  14  may have a diameter of between approximately 6.0 mm and approximately 7.0 mm, and a height of between approximately 10.0 mm and approximately 11.0 mm. 
     Another embodiment of sensor mount  18  can be seen in  FIG. 7 , in which a sensor  68  is secured to a side surface of sensor mount  18 . In another embodiment, as seen in  FIG. 8 , sensor  68  could be secured to a bottom surface of sensor mount  18 . It is to be appreciated that in the embodiment of  FIG. 8 , sensor mount  18  could have a cylindrical shape, and sensor  68  could have a circular shape. 
     Sensor  68  may be, in certain embodiments, a graphene or a carbon nanotube sensor, e.g., a functionalized graphene or carbon nanotube substrate. Such a graphene or a carbon nanotube sensor can detect changes in conductance when a target analyte or plurality of target analytes contact the functionalized graphene or carbon nanotube substrate. 
     In certain embodiments, such a carbon nanotube substrate includes semiconducting single walled carbon nanotubes (s-SWCNTs). Such s-SWCNTs are characterized by a high surface area and semiconducting properties sufficient to produce a scalable sensitivity. In certain embodiments the carbon nanotube substrate may be planar. The carbon nanotube substrate may be a carbon nanotube semiconductor surface fashioned into a biosensor device that monitors electrical field charge carriers across the semiconductor material&#39;s surface. When binding events from biomolecular interactions occur and are coupled with the surface of the carbon nanotubes, the carrier concentration on the nanotube can change, which changes the conductivity. As target analytes bind to the functionalized nanotube surface, the current is altered and detected. In certain embodiments, the binding interaction occurs within the Debye screening length in order for the interaction to be detected. To enhance the sensitivity, small receptors such as fragmented antibodies, can be used. 
     The carbon nanotubes can be single walled carbon nanotubes known to those of skill in the art and generally used for the manufacture of carbon nanotube substrates. Carbon nanotubes (CNTs), as are known in the art, are allotropes of carbon with a generally cylindrical nanostructure. In general, carbon nanotubes are characterized by a hollow cylindrical structure of given length with the walls formed by one-atom-thick sheets of carbon, called graphene. In general, graphene sheets are rolled or otherwise configured at specific and discrete (“chiral”) angles, and the combination of the rolling angle and radius decides the nanotube properties, for example, whether the individual nanotube shell is a metal or semiconductor. Nanotubes are categorized as single-walled nanotubes (SWCNTs) and multi-walled nanotubes (MWCNTs). Individual nanotubes can naturally align themselves into “ropes” held together by van der Waals forces, more specifically, pi-stacking. Exemplary single-walled carbon nanotubes (SWCNTs) have a diameter of about 1-2 nanometer, but can be wider. According to one aspect, SWCNTs can exhibit a band gap from zero to about 2 eV and their electrical conductivity can show metallic or semiconducting behavior. Single-walled carbon nanotubes provide exemplary substrates for the detection devices described herein. A more detailed description of exemplary single-walled carbon nanotubes and methods of their manufacture are described in U.S. application Ser. No. 16/155,955, filed on Oct. 10, 2018, and entitled “Carbon Nanotube-Based Device for Sensing Molecular Interaction,” the entire disclosure of which is incorporated herein by reference in its entirety for all purposes. Exemplary carbon nanotubes for use in devices are also described in U.S. Pat. Nos. 7,416,699, 6,528,020, and 7,166,325 each of which is hereby incorporated by reference in its entirety. 
     Those having skill in the art, with the knowledge gained from the present disclosure, will recognize that various changes can be made to the disclosed apparatuses and methods in attaining these and other advantages, without departing from the scope of the present invention. As such, it should be understood that the features described herein are susceptible to modification, alteration, changes, or substitution. For example, it is expressly intended that all combinations of those elements and/or steps which perform substantially the same function, in substantially the same way, to achieve the same results are within the scope of the invention. Substitutions of elements from one described embodiment to another are also fully intended and contemplated. The specific embodiments illustrated and described herein are for illustrative purposes only, and not limiting of the invention as set forth in the appended claims. Other embodiments will be evident to those of skill in the art. It should be understood that the foregoing description is provided for clarity only and is merely exemplary. The spirit and scope of the present invention are not limited to the above examples, but are encompassed by the following claims. All publications and patent applications cited above are incorporated by reference in their entirety for all purposes to the same extent as if each individual publication or patent application were specifically and individually indicated to be so incorporated by reference.