Patent Publication Number: US-2005118707-A1

Title: Positioning system for moving a selected station of a holding plate to a predetermined location for interaction with a probe

Description:
The present application is a continuation-in-part of pending U.S. patent application Ser. No. 09/894,956 filed Jun. 27, 2001, which is a continuation-in-part of pending U.S. patent application Ser. No. 09/687,219, filed Oct. 12, 2000, which is a continuation-in-part of pending U.S. patent application Ser. No. 09/444,112, filed Nov. 22, 1999, which is a continuation-in-part of pending U.S. patent application Ser. No. 08/876,276, filed Jun. 16, 1997; additionally, the present application is a continuation-in-part of pending U.S. patent application Ser. No. 09/636,778, filed Aug. 11, 2000, which application is a continuation and claims the benefit of priority under 35 U.S.C. § 120 of U.S. patent application Ser. No. 09/098,206, filed Jun. 16, 1998, which issued as U.S. Pat. No. 6,174,673 on Jan. 16, 2001, which is a continuation-in-part of pending U.S. patent application Ser. No. 08/876,276, filed Jun. 16, 1997, all of the contents of which are incorporated by reference in their entirety herein. 
    
    
     FIELD OF THE INVENTION  
      The present invention pertains generally to devices for performing operations on selected samples in a holding plate with a probe. More particularly, the present invention pertains to positioning systems for moving a selected station of a holding plate to a predetermined location for interaction with a p robe. The present invention is particularly, but not exclusively, useful as a computer assisted, optical system for positioning a holding plate having over a thousand, small diameter through-hole stations at a precise location to allow a probe to interact with a selected station.  
     BACKGROUND OF THE INVENTION  
      Plates for holding specimen samples in a fluid solution are available having over a thousand, small diameter stations. The stations can include through-holes, or wells that extend only partially into the holding plate. In the case of a through-hole station, these stations rely on surface tension to hold each fluid sample in a respective station. The through-hole stations of a holding plate can be filled with a solution of interest by simply immersing a surface of the holding plate into the solution. Capillary action causes the solution to enter the through-hole stations. This allows a very large number of relatively small volume samples of the solution to be simultaneously prepared for later analysis or manipulation. Specifically, holding plates having over a thousand stations arranged in a planar array, with station diameters of only about 500 microns, are available.  
      Once the holding plate has been filled with solution, it is often desirable to either add material to selected stations or withdraw solution from selected stations. This is particularly the case when the solution used to fill the holding plate is non-homogenous. Often times, the selected stations differ in color, opacity, fluorescence or are otherwise optically distinguishable from the remaining stations. For example, a biological or chemical reaction may proceed more rapidly in portions of the solution, causing only selected stations to change color, while the remaining stations do not. Withdrawal of solution from the selected stations allows for the separation of the solution into portions of solution that have reacted and portions of solution that have not reacted. Alternatively, it may be desirable to add a material such as a chemical reagent to selected stations, again selecting stations based on some optical property of the sample in the station.  
      Generally, a thin, needle-like probe must be positioned in fluid communication with a selected station to either add or withdraw material from the station. Thus, it is often desirable to select a specific station based on an optical characteristic of the station&#39;s sample and then operate on the selected station with a probe. To accomplish this, the probe and selected station must first be aligned. Unfortunately, for stations having extremely small diameters, such as through-holes with diameters of 500 microns or less, it is impossible for all practical purposes, to manually align a selected station with a probe. Thus, the present invention recognizes that a computer-assisted, automated system is necessary to align small diameter stations with a probe.  
      Holding plates are generally designed with stations (i.e. through-holes or wells) having station axes that are perpendicular to the sides of the holding plate. With this design, the axes of the stations are relatively easily aligned with the path of the probe. Unfortunately, due to defects in the manufacturing processes that are used to prepare the holding plates, the axes of the stations can sometimes be misaligned, albeit slightly, from the sides of the holding plate. Stated another way, an end of the station on one side of a holding plate is offset from the other end of the station on the opposite side of the holding plate. It is to be appreciated that this offset can p resent problems when imaging is performed on one side of the holding plate while the probe is aligned with the station on the opposite side of the holding plate. The problem becomes more egregious with respective increases in the aspect ratio of the station, the density of stations on the plate and the thickness of the plate.  
      It is often the case that hundreds of stations (among the thousand or more stations present in the holding plate) may require interaction with the probe. In these cases, it becomes too labor intensive for an operator to select each station individually for interaction with the probe. Thus, it would be desirable to have a computer-assisted system that allows the operator to select a s et of stations by merely choosing an optical characteristic to establish the set. With the set established, the operator then instructs the computer to successively perform a probe operation on each station in the selected set. A convenient system would allow an operator to specify an optical characteristic, for example—fluorescence, and then instruct the computer to make a chemical addition to each station having a green sample that is fluorescing.  
      In light of the above, it is an object of the present invention to provide a system suitable for the purposes of moving a selected station of a holding plate to a predetermined location for interaction with a probe. It is another object of the present invention to provide a positioning system for aligning a probe and selected station wherein the station has an extremely small diameter (i.e. a through-hole having a diameter of 500 microns or less). It is yet another object of the present invention to provide a system for automatically performing a probe operation on samples in a selected set of stations that all have a common optical characteristic. Still another object of the present invention is to provide a positioning system for aligning a probe with a selected station wherein the station axis is offset (i.e. at a non-normal angle) relative to the side of the holding plate. Yet another object of the present invention is to provide an optical positioning system for aligning a small diameter through-hole station with a probe which is efficient to use, relatively simple to implement, and comparatively cost effective.  
     SUMMARY OF THE INVENTION  
      The present invention is directed to a device for positioning the tip of a probe at a selected station of a holding plate. For the present invention, the holding plate is formed with a substantially flat first side and an opposed second side. Preferably, the holding plate is further formed with a regular or irregular planar array of stations for holding a plurality of respective samples. Importantly, each station is accessible by the probe from the first side of the holding plate.  
      In accordance with the present invention, the probe is attached to a base and a mechanism is provided to allow for reciprocal movement of the probe relative to the base. The device further includes a moveable stage that is mounted on the base to support the holding plate. For the present invention, the moveable stage is formed with a planar surface for engagement with the second side of the holding plate. With this cooperation of structure, the planar surface of the stage defines a coordinate plane (m xy ) containing orthogonal axes x and y. A mechanism is provided to secure the holding plate to the stage, causing the holding plate to move with the stage. With the second side of the holding plate secured against the stage, the first side of the holding plate remains exposed for interaction with the probe. To selectively move the stage (and the holding plate) in the x and y directions relative to the base and probe, the device further includes a pair of motorized linear actuators.  
      As indicated above, the probe is attached to the base. In greater structural detail, the probe is elongated and defines a probe axis in the direction of elongation. For the present invention, the elongated probe is optically distinguishable and, for this purpose, is preferably mounted on a fluorescent hub and extends from the fluorescent hub to a probe tip. The hub, in turn, is mounted on the base. Importantly for the present invention, the probe is positioned relative to the holding plate to allow the tip of the probe to interact with the first side of the holding plate. Additionally, the probe and hub are preferably mounted on the base with the probe axis of the probe oriented normal to the m xy  plane. In the preferred embodiment of the present invention, a mechanism is provided to allow the probe to reciprocate (relative to the holding plate and base) along the probe axis and in a direction that is substantially orthogonal to the m xy  plane. With the above described combination of structure, the motorized linear actuators can be used to move the holding plate to a location in the m xy  plane such that a selected station is positioned on the probe axis. With the selected station positioned on the probe axis, the probe can be moved along the probe axis to interact with the selected station.  
      To locate a selected station of the holding plate at a position on the probe axis, the device includes at least one camera and a computer processor. In the preferred embodiment of the present invention, the camera is positioned on the probe axis and oriented to obtain a pixel image of the holding plate stations from the second side of the holding plate. To facilitate imaging from the second side of the holding plate, a transparent stage is preferably used. Alternatively, one or more holes can be formed in the stage to allow the camera to image the stations from the second side of the holding plate.  
      In operation, the device is initially calibrated (calibration procedure described below). Next, a first holding plate is installed on the stage, placing the holding plate at a first location in the m xy  plane. The expectation at this point is that there will be an optical contrast between various stations in the holding plate. One or more pixel images are then obtained by the camera that images the array of stations positioned at the first location in the m xy  plane and the projection of the probe in the m xy  plane. For the present invention, the pixel image defines a coordinate plane (p xy ) that is related to the coordinate plane (m xy ). From the pixel image, the operator selects a specific station of the holding plate that requires interaction with the probe. This information is then transferred to t he computer processor. The computer processor instructs the motorized linear actuators to move the holding plate through the proper x and y distances in the m xy  plane to align the selected station on the probe axis. More specifically, the computer uses a relationship that was previously established between the coordinate plane (p xy ) and the coordinate plane (m xy ) during calibration to accurately move the stage and align the selected station on the probe axis. With the selected station positioned on the probe axis, the probe is then translated along the probe axis to interact with the station. In one embodiment of the present invention, station offset information (i.e. the deviation of each station axis from a reference axis that is orthogonal to the side of the holding plate) is input into the computer processor. The computer processor then uses the offset information to ensure that the station entrance located at the first side of the holding plate is aligned with the probe axis.  
      To calibrate the device, an optical marker is placed on the stage and a first pixel image is obtained by the camera. As such, the first pixel image includes the optical marker positioned at a first location in the m xy  plane. Preferably, the calibration procedure is performed without a holding plate on the stage. Next, the stage is moved using the motorized linear actuators to successive locations in the m xy  plane. The actuator displacements (e.g. motor steps) necessary to move the optical marker between locations are recorded and a pixel image of the optical marker is obtained at each location. These pixel images and actuator displacements are then used by the computer processor to correspond the p xy  coordinate plane with the m xy  coordinate plane. Stated another way, the pixel images are used to find the relationship between the p xy  coordinate plane and the m xy  coordinate plane. Preferably, the method of least squares is used to establish an approximate linear relationship between the coordinate plane (p xy ) and the coordinate plane (m xy ).  
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
      The novel features of this invention, as well as the invention itself, both as to its structure and its operation, will be best understood from the accompanying drawings, taken in conjunction with the accompanying description, in which similar reference characters refer to similar parts, and in which:  
       FIG. 1  is a perspective view of a device in accordance with the present invention for moving a selected station of a holding plate to a predetermined location for interaction with a probe;  
       FIG. 2  is an enlarged, sectional view of a portion of a holding plate and stage as would be seen along line  2 - 2  in  FIG. 1 ;  
       FIG. 3A  is an exemplary pixel image taken after the optical marker has been moved to a first location;  
       FIG. 3B  is an exemplary pixel image taken after the optical marker has been moved to a second location;  
       FIG. 3C  is an exemplary pixel image taken after the optical marker has been moved to a third location; and  
       FIG. 4  is a sectional view as in  FIG. 2  showing a holding plate with offset stations.  
    
    
     DESCRIPTION OF THE PREFERRED EMBODIMENT  
      Referring initially to  FIG. 1 , a system  10  for performing operations on selected samples in a holding plate  12  with a probe  14  in accordance with the present invention is shown. As shown, the system  10  includes a base  16  for supporting both the holding plate  12  and the probe  14 . As further shown, the probe  14  is preferably elongated and defines a probe axis  18  in the direction of elongation. In the preferred embodiment of the present invention, the probe  14  is formed as a hollow needle having a lumen capable of transferring fluid. Also shown in  FIG. 1 , the elongated probe  14  is preferably mounted on a hub  20  and extends from the hub  20  to a probe tip  22 . For the present invention, the hub  20 , which is preferably fluorescent, is somehow optically distinguishable from the probe  14 . The system  10  also includes a mechanism  24  to move the probe  14  back and forth along the probe axis  18 , relative to the base  16  and holding plate  12 . Those skilled in the art will appreciate that any mechanism  24  known in the pertinent art for reciprocating a probe back and forth along an axis, such as a hydraulic or pneumatic cylinder, can be used in conjunction with the present invention.  
      With cross reference now to  FIGS. 1 and 2 , it can be seen that the holding plate  12  is formed with a substantially flat first side  26  and an opposed second side  28 . Preferably, the holding plate  12  is further formed with a regular or irregular planar array of stations  30 , for which stations  30   a - c  shown in  FIG. 2  are exemplary. Each station  30  is provided to hold a fluid sample and may optionally be a through-hole that extends through the plate  12  between sides  26  and  28 . In the preferred embodiment of the present invention, the holding plate  12  is formed with over one thousand stations  30 , with each station  30  having an inner diameter  31  of approximately 500 microns or less. An optional coating  32  can be applied to each through-hole station  30  to limit the transmission of light between adjacent stations  30 . Importantly, each station  30  is accessible by the probe  14  from the first side  26  of the holding plate  12 .  
      With continued cross reference to  FIGS. 1 and 2 , it can be seen that the system  10  further includes a moveable stage  34  that is mounted on the base  16  to support the holding plate  12 . As further shown, the moveable stage  34  is formed with a planar surface  36  for engagement with the second side  28  of the holding plate  12 . As shown, the planar surface  36  of the stage  34  defines a coordinate plane (m xy ) containing orthogonal axes x and y. If required, clamps (not shown) can be provided to secure the holding plate  12  to the stage  34 . In any case, with the holding plate  12  on the stage  34 , the stage  34  and holding plate  12  move together. With the second side  28  of the holding plate  12  secured against the stage  34 , the first side  26  of the holding plate  12  remains exposed for interaction with the probe  14 .  
      As best seen in  FIG. 1 , the system  10  includes a pair of motorized linear actuators  38   a, b  that are mounted on the base  16  to selectively move the stage  34  and holding plate  12  in the x and y directions relative to the base  16  and probe  14 . It is to be further appreciated that the motorized linear actuators  38   a, b  move the holding plate  12  within the m xy  plane. Preferably, each motorized linear actuator  38   a, b  includes a stepper motor for driving a lead screw to move the stage  34 . For the present invention, any type or number of motorized linear actuators or other devices known in the pertinent art for selectively moving a stage in at least two directions can be used.  
      Referring now with cross reference to  FIGS. 1 and 2 , it can be seen that the probe  14  is positioned relative to the holding plate  12  to allow the probe tip  22  to interact with the first side  26  of the holding plate  12 . Additionally, the probe  14  is preferably mounted on the base  16  with the probe axis  18  of the probe  14  oriented normal to the m xy  plane (i.e. the plane containing the x and y axes). Thus, the probe  14  reciprocates along the probe axis  18  and in a direction that is orthogonal to the m xy  plane. In accordance with the present invention, the motorized linear actuators  38   a, b  can be selectively activated to move the holding plate  12  to a location in the m xy  plane such that a selected station  30  is positioned on the probe axis  18 . With the selected station  30  positioned on the probe axis  18 , the probe  14  can then be moved along the probe axis  18  to interact with the selected station  30 . More specifically, the probe  14  can manipulate a sample that is held by the holding plate  12  at the selected station  30 . Manipulations of the sample by the probe  14  can include sample withdrawal from the station  30  or the addition of a material such as a chemical reagent to the sample.  
      As best seen in  FIG. 1 , the system  10  includes a camera  40  and a computer processor  42  with a display  44 . Preferably, as shown, the camera  40  is positioned on the probe axis  18  and oriented to image the stations  30  of the holding plate  12  from the second side  28  (shown in  FIG. 2 ) of the holding plate  12 . The camera  40  produces a pixel image  46  that can be displayed on the display  44 . The holding plate  12  can be imaged through transparent portions of the stage  34  and base  16 , or one or more holes can be formed in the stage  34  and base  16 .  
      In the preferred embodiment of the present invention, the system  10  further includes an illumination system  48  for illuminating and/or exciting samples in the holding plate  12 . For example, the illumination system  48  can be used to excite fluorescent materials in the holding plate  12 . In accordance with the present invention, one or more light filters  50  can be used to selectively filter light entering the camera  40 . For example, light filter  50  can be used to filter out backscattered excitation light from illumination system  48  while allowing fluorescent emissions from the samples to be imaged by the camera  40 .  
      In operation, a holding plate  12  is installed on the stage  34 , as shown in  FIG. 1  and a pixel image  46  is created by camera  40  and presented in a viewable format by display  44 . As shown, the pixel image  46  sequentially includes a hub image  52 , a probe image  54  and an image of the array of stations  30  of the holding plate  12 . In part, because the probe  14  is surrounded by an optically distinguishable hub  20 , the relatively thin probe  14  can be imaged. It is to be appreciated that the pixel image  46  also shows stations  30 , including stations  30  that have distinguishing optical characteristics (e.g. color, fluorescence, opacity, etc). In  FIG. 1 , pixel image  46  shows the image of five selected stations  30  that have distinguishing optical characteristics (i.e. selected stations image  56 ).  
      As indicated above, the function of the system  10  is to move the holding plate  12  within the m xy  plane to position a selected station  30  on the probe axis  18 . With the selected station  30  on the probe axis  18 , the probe  14  is then moved along the probe axis  18  to manipulate a sample in the selected station  30 . For the present invention, the pixel image  46  defines a coordinate plane (p xy ) that is related to the coordinate plane (m xy ). In accordance with the present invention, stations  30  are selected in the pixel image  46  for manipulation by the probe  14 . The computer processor  42  then instructs the motorized linear actuators  38   a, b  to move the holding plate  12  within the m xy  plane to position the selected station  30  on the probe axis  18 . In accordance with the present invention, the system  10  is calibrated to accomplish this movement with extremely small positional errors. During calibration, the computer processor  42  determines the relationship (i.e. correspondence) between the coordinate plane (p xy ) and the coordinate plane (m xy ).  
      To establish the relationship between the coordinate plane (p xy ) and the coordinate plane (m xy ), an optical marker is placed on the stage  34  and the stage  34  is moved via the motorized linear actuators  38   a, b  to successive locations in the m xy  plane. A separate pixel image  46  is obtained at each location. The displacements of the motorized linear actuators  38   a, b  (e.g. motor steps) necessary to move the optical marker from the first location to the second location and from the second location to the third location are recorded and input into the processor  42 .  
       FIGS. 3A, 3B  and  3 C show pixel images  46 ′,  46 ″ and  46 ′″ for three locations of the stage  34  within the m xy  plane. In greater detail,  FIG. 3A  shows pixel image  46 ′ for stage  34  in a first location and includes an optical marker image  58 ′. Similarly,  FIG. 3B  shows pixel image  46 ″ for stage  34  in a second location and includes an optical marker image  58 ″. Also,  FIG. 3C  shows pixel image  46 ′″ for stage  34  in a third location and includes an optical marker image  58 ′″. Although pixel images  46 ′,  46 ″ and  46 ′″ for three stage  34  locations are shown herein, it is to be appreciated that any number of locations can be used with the present invention to establish a relationship between the coordinate plane (p xy ) and the coordinate plane (m xy ). Once the displacements of the motorized linear actuators  38   a, b  (e.g. motor steps) and pixel images  46 ′,  46 ″ and  46 ′″ have been obtained, a linear regression technique, such as the method of least squares, can be used by the processor  42  to establish an approximate linear relationship between the coordinate plane (p xy ) and the coordinate plane (m xy ) to calibrate the system  10 .  
      Referring now to  FIG. 4 , a portion of a holding plate  12  having a thickness, “t”, is shown. The holding plate  12  includes a station  30  with a station entrance (top)  60  that is offset from the station exit (bottom)  62 . As further shown, the axis  64  of the station  30  is inclined at an angle, α, from an axis  66 . More specifically, the axis  66  is normal to the side  26  of the holding plate  12  and passes through the exit (bottom)  62 . It can be further seen that a line  67  on side  26 , which intersects both the axis  66  and the axis  64  establishes a rotation angle, θ, between the line  67  and a base reference line  68  about the axis  66 . In one embodiment of the present invention, this offset information (i.e. α, θ, and “t”) for the plate  12  is input into the computer processor  42 . With this offset information, the computer processor  42  uses an image of the second side  28  of the plate  12  to accurately locate the entrance  60  of the plate  12  on the probe axis  18  (probe axis  18  shown in  FIG. 1 ).  
      While the particular positioning system for moving a selected station of a holding plate to a predetermined location for interaction with a probe as herein shown and disclosed in detail is fully capable of obtaining the objects and providing the advantages herein before stated, it is to be understood that it is merely illustrative of the presently preferred embodiments of the invention and that no limitations are intended to the details of construction or design herein shown other than as described in the appended claims.