Abstract:
A system for depositing molecular liquids on a receiver comprising: a printing station having one or more print heads spanning the width of a receiver to be printed on; a receiver transport mechanism for transporting a receiver through the printing station so that the one or more print heads can deposit molecular liquids in an array on the receiver; a maintenance and service station located in proximity to the printing station; and a printhead translation mechanism for moving a printhead to the maintenance and service station to receive maintenance and service.

Description:
FIELD OF THE INVENTION  
         [0001]    This invention relates in general to molecular biological systems and, more particularly to a means by which micro-array receivers of molecular biological reagents and samples can be produced. More particularly, the invention provides a means by which small volumes of molecular biological liquids can be deposited onto rigid, semi-rigid or flexible supports for the production of micro-array receivers.  
         BACKGROUND OF THE INVENTION  
         [0002]    As is well known (and described for example in U.S. Pat. No. 5,807,522, inventors Brown et al. and in “DNA Microarrays: A Practical Approach”, Schena, Mark, New York, Oxford University Press, 1999, ISBN 0-19-963776-8), micro-arrays are arrays of very small samples of purified DNA or protein target material arranged as a grid of hundreds or thousands of small spots on a substrate. When the micro-array is exposed to selected probe material, the probe material selectively binds to the target spots only where complementary bonding sites occur, through a process called hybridization. Subsequent quantitative scanning in a fluorescent micro-array scanner may be used to produce a pixel map of fluorescent intensities (See, e.g., U.S. Pat. No. 5,895,915, inventors DeWeerd et al.). This fluorescent intensity map can then be analyzed by special purpose algorithms that reveal the relative concentrations of the fluorescent probes and hence the level of gene expression, protein concentration, etc., present in the cells from which the probe samples were extracted.  
           [0003]    Historically, microarrays could be constructed either manually or mechanically through the use of photolithographic, robotically controlled or other apparatus for the precise metering and placement of molecules. Alternatively, microarrays could be constructed through direct chemical synthesis on a solid support. Such devices and methods have the undesirable result that micro-arrays with a great number of individual spots and thus a great number of individual molecular biological reagents are contained with little or no means to identify them uniquely, either by human observations or machine.  
           [0004]    Many examples exist for dispensing liquids in small volumes in the range of milliliters to sub-fractions of milliliters. For example, Pastinen et al. (Genorne Research, 7-606-614 (1997)) create an array of oligonucleotides by manually applying 0.5˜IL of a solution of 5′-amino-modified oligonucleotides onto an epoxide-activated glass slide to produce a 3×3 array of oligonucleotides on a 0.36 cm˜ area of a preprinted glass slide.  
           [0005]    Other, more traditional printing methods have been used to create patterns of a few different reagents on a solid support. Means such as silk screening, offset printing, and rotogravure printing have been used in the production of reagent test strips. In such methods, each reagent ink is applied separately. Johnson, for example, (U.S. Pat. No. 4,216,245) discusses methods for the production of reagent test strip devices.  
           [0006]    Pipette dispensing of reagents can be automated. Automation potentially increases the speed and accuracy of array production, while decreasing the necessary spacing between array positions. However, the utility of automated pipetting methods are severely limited in the number of different reagents that may be simultaneously applied (low parallelism). Cozzette et al., for example, (U.S. Pat. No. 5,554,339) discusses the use of microsyringes for dispensing reagents during the production of bio-sensor devices.  
           [0007]    High-speed robotics have also been used to print micro-arrays of amino-modified cDNA molecules onto silylated glass microscope slides (CEL Associates, Houston) or poly-l-lysine coated microscope slides (Schena, BioEssays, 18:427-431 (1996); Schena et al., Proc. Nati. Acad. Sci., U.S.A., 93:10614-10619 (1996).  
           [0008]    Another approach to microarray printing is an adaptation of inkjetting technology. For example, Hayes et al., U.S. Pat. No. 4,877,745 discusses an ink-jet type method and apparatus for dispensing reagents, particularly in the production of reagent test strips.  
           [0009]    Pin transfer is one approach that allows the simultaneous transfer of greater numbers of samples than possible with the above approaches. Examples of such pins are discussed in U.S. Pat. No. 5,770,151, inventors Roach et al. and U.S. Pat. No. 5,807,522, inventors Brown et al.  
           [0010]    Pirrung et al., U.S. Pat. No. 5,143,854, Fodor et al., U.S. Pat. No. 5,510,270, inventors, Fodor et al., U.S. Pat. No. 5,445,934, and Chee et al., International Patent Application, WO 95/11995 discuss the production of high  2  density oligonucleotide arrays through a photolithographic, directly onto a derivatized glass substrate.  
           [0011]    McGall et al., U.S. Pat. No. 5,412,087 discusses a method for the production of a high density oligonucleotide array from pre-sythesized oligonucleotides.  
           [0012]    Birch et al, U.S. Pat. No. 6,051,190 and U.S. Pat. No. 6,303,387 discusses a transfer rod for distribution of small amounts of liquid in biological or chemical analysis.  
           [0013]    Bryning et al, U.S. Pat. No. 6,296,702 BI discusses an oscillating fiber apparatus for dispensing small volumes of a selected liquid onto a substrate. Similarly, Dannoux et al, International Patent Application WO 00/30754 discusses a method and apparatus for printing high-density biological arrays utilizing a plurality of rods housed with a channel.  
           [0014]    Capillary transfer is another approach that allows the simultaneous transfer of greater numbers of samples. Chen et al, US Patent Application Publication No. 2001/0053334 discusses a print system and method of printing probe micro-arrays with capillary bundles. Similarly, Rogers et al., WO 00/01859 discusses a gene pen apparatus for repetitive printing of arrays.  
           [0015]    In view of the above, the need is apparent for an efficient system for depositing molecular biological reagents and samples that are contained on solid or semi-solid or flexible supports.  
         SUMMARY OF THE INVENTION  
         [0016]    According to the present invention, there is provided a solution to the problems discussed above.  
           [0017]    According to a feature of the present invention, there is provided a system for depositing molecular liquids on a receiver comprising:  
           [0018]    a printing station having one or more print heads spanning the width of a receiver to be printed on;  
           [0019]    a receiver transport mechanism for transporting a receiver through said printing station so that said one or more print heads can deposit molecular liquids in an array on said receiver;  
           [0020]    a maintenance and service station located in proximity to said printing station; and  
           [0021]    a printhead translation mechanism for moving a printhead to said maintenance and service station to receive maintenance and service.  
         ADVANTAGEOUS EFFECT OF THE INVENTION  
         [0022]    The invention has the following advantages.  
           [0023]    1. Improved systems productivity is provided for the high speed production of microarrays of biological and chemical molecules on a rigid, semi-rigid or flexible supports.  
           [0024]    2. A system is provided for depositing a large number of unique small volumes of molecular biological and chemical liquids on a substrate.  
           [0025]    3. A system is provided wherein printheads can be easily removed, maintained and serviced. 
       
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0026]    [0026]FIG. 1 is a diagrammatic view showing the micro-deposition system for biomolecular fluids, including the printing and maintenance and service regions.  
         [0027]    [0027]FIG. 2 is a diagrammatic view showing the “Net-shaped printhead with fluid connections and ejectors fluidically coupled to the supply lines.  
         [0028]    [0028]FIG. 3 diagrammatically shows the elements contained in the maintenance and service region.  
         [0029]    [0029]FIG. 4 diagrammatically shows the printhead engaged with a maintenance element.  
         [0030]    [0030]FIG. 5 diagrammatically shows the relative movement of the printhead and the maintenance element including a compliant member that keeps the maintenance element in intimate contact with the printhead.  
         [0031]    [0031]FIG. 6 diagrammatically shows a maintenance element with vacuum to pull fluids into the printhead assembly and to provide service of the printhead by vacuum assistance. A recessed area is included to provide a region for the vacuumed fluid to reside away from the printhead surface.  
         [0032]    [0032]FIG. 7 diagrammatically shows a maintenance element in which an absorbing material is included.  
         [0033]    [0033]FIG. 8 diagrammatically shows a maintenance element wherein a wiper blade is incorporated.  
         [0034]    [0034]FIG. 9 diagrammatically shows a patterned array and magnification of a sub-region of the array created by the invention.  
         [0035]    [0035]FIG. 10 diagrammatically shows a detailed view of the sub-region of the patterned array shown in FIG. 9.  
         [0036]    [0036]FIG. 11 diagrammatically shows a detailed view of the array as shown in FIG. 9.  
         [0037]    [0037]FIG. 12 diagrammatically shows a detailed view of a droplet ejected from the printhead to create the sub-array.  
         [0038]    [0038]FIG. 13 diagrammatically shows a detailed view of a macro droplet ejected from the printhead of sufficient fluid volume to cover the entire region of the sub-array.  
         [0039]    [0039]FIG. 14 is a diagrammatic view showing a “Net Shaped” piezoelectric material with cylindrical void.  
         [0040]    [0040]FIG. 15 is a diagrammatic view showing an electrode configuration for the “Net Shaped” material of FIG. 14.  
         [0041]    [0041]FIG. 16 is a diagrammatic view showing a simple print head configuration with a glass capillary tube inserted into the void and a fluidic connection.  
         [0042]    [0042]FIG. 17 is a diagrammatic view showing a print head configuration of FIG. 3 with an orifice plate attached to the glass capillaries.  
         [0043]    [0043]FIG. 18 is a diagrammatic view showing a print head configuration of a linear array of capillary tubes.  
         [0044]    [0044]FIG. 19 is a diagrammatic view showing a print head configuration of a matrix of capillary tubes.  
         [0045]    [0045]FIG. 20 is a diagrammatic view of a print head configuration where the voids created by the “Net shaped” process is the channel for molecular biological liquids. 
     
    
     DETAILED DESCRIPTION OF THE INVENTION  
       [0046]    In general, this invention relates to a system using a print head devices for micro-deposition of molecular biological or chemical liquids on a solid or semi-solid or flexible support. Approximately 1000 molecular biological liquids need to be uniquely placed on a 2-D grid, each solution occupying approximately 50-500 micro meter (um) diameter spot and preferably 50-200 um spot diameter. This invention is advantaged in that it provides an efficient means by which a large number of small volume molecular biological reagents can be deposited.  
         [0047]    In the following description, a preferred print head will be described with reference to FIGS.  14 - 20 , and then a system utilizing a plurality of such printheads will be described with reference to FIGS.  1 - 13 .  
         [0048]    Specifically, a print head is proposed where the deposition process is created by a pressure pulse derived from a piezoelectric element. This element is constructed by a process know as “net shaping” as discussed in Chatterjee et al., U.S. Pat. Nos. 6,065,195 and 6,168,746. This process provides the advantage of producing complex 3-D (three-dimensional) mechanical shapes with reduced manufacturing steps. As discussed in U.S. Pat. No. 6,168,746, this process consists of the steps: spray drying fine particulate ceramic ferroelectric material to form agglomerate material; mixing the spray dried fine particulate ceramic ferroelectric agglomerate material with a binder system including materials selected from the group consisting of wax having wax components of different molecular weight, magnesium-X silicate, agaroid gel forming material, and agaroid gel forming material mixed with magnesium-X silicate to form a compounded material; injecting the compounded material at a selected pressure into a mold to form a green article; debinding or drying the green article; sintering the debinded or dried green article to form the final molded article; poling the final molded article to align the electrical dipoles within the piezoelectric material; and forming a coating of conductive material over the top and bottom surfaces of the final molded article.  
         [0049]    As shown in FIG. 14, a block  10  of ferroelectric material, preferably a piezoelectric material and preferably lead zirconate titinate (PbZrTiO 3 ) is formed to create a geometry with cylindrical voids  12 . A first electrode  20  (FIG. 15) covers void  12  and a second electrode  22  covers block  10 . The poling process is done such that when a voltage is applied between electrodes  20 , 22 , a radial force is created at the cylindrical void  12 . As shown in FIG. 16, each void contains a glass or plastic capillary  30  that is held in place with suitable cement. Examples of glass capillaries suitable for this application are available from Nippon Electric Glass, Inc. Capillary inside diameters on the order of 30-100 um and preferably in the range of 30-60 um are appropriate. The aforementioned radial force acts on the tube, which contains the molecular biological liquids, ejecting a drop of known volume. The molecular biological or chemical liquids are connected to the glass capillaries via suitable flexible or rigid tubing  32 . A variant of this embodiment is shown in FIG. 17 includes an orifice plate  40  having orifices  42  that would cover the ends of the glass capillary(s).  
         [0050]    In yet another variant of this embodiment shown in FIG. 18, the piezoelectric element contains a linear array of 1×N capillary elements  52 .  
         [0051]    Yet another embodiment shown in FIG. 19, the piezoelectric element  60  contains an M×N array of capillary elements  62 .  
         [0052]    In another embodiment of this invention shown in FIG. 20, a block of ferroelectric material  70 , preferably a piezoelectric material and preferably lead zirconate titinate (PbZrTiO 3 ) is formed to create a molded geometry with cylindrical voids  72  where each void is the channel for containing molecular biological and chemical liquids. An orifice plate  74  with apertures  76  covers the end of the molded channels. The shape of the voids could be geometries other than circular such as square or rectangular.  
         [0053]    An electric signal is applied to the electrodes (See FIG. 2) to produce the necessary force to produce the ejection of a drop of liquid.  
         [0054]    A system for producing a receiver (support) containing bio-specific solutions is described with reference to FIGS.  1 - 13 . The system contains a printing station  90  1 or more “Net-Shaped” page-width printheads  100 - 108 , a fluid delivery system  110 ,  112 , printhead service/maintenance station and a receiver transport mechanism  116  and printhead translation mechanism  118 . Computer  111  controls mechanisms  116 , 118 , fluid deposition  110 , 112  and maintenance and service station  114 . Approximately 1000 molecular biological liquids need to be uniquely placed on a 2-D grid, each solution occupying approximately 50-500 micro meter (um) diameter spot and preferably 50-200 um spot diameter. This invention is advantaged in that it provides for an efficient means to effectively produce arrays of biomolecules on a support, a means to easily remove printheads in the event they require service and a means for high-throughput array generation.  
         [0055]    Specifically, a system is provided where bio-specific solutions can be efficiently placed at known locations. In one embodiment of this invention, a “Net-Shaped” page-width print head bar is described wherein the spacing of the bio-specific solutions on the support are matched equally to the spacing of the printhead nozzles. In addition, a system where the printhead nozzles are not equally spaced with respect to the dots formed on the support envisioned and is accomplished by the combination of printhead and receiver motion that is coupled to provide the desired dot spacing.  
         [0056]    The printing or deposition of these sites would be created in the Printing Station  90  as shown in FIG. 1. The advantage of the system described in this application and shown in FIG. 1 is the ability to move the printheads in a direction  120  normal to the direction  122  of printing. This permits 2 features: 1) the ability to create unique patterns of bio-specific sites, and 2) the ability to move the “Net-shaped” printhead (shown in FIG. 2) to a Maintenance and Service Station  114  with maintenance elements as described in FIG. 3 and shown in detail in FIG. 4 and more specifically in FIG. 5 where the printhead motion relative to the maintenance element is shown. FIG. 2 shows printhead  100  as including two rows of nozzles  130  supplied by fluid line  132 . FIG. 3 shows maintenance and service station  114  includes maintenance elements  134  aligned with print heads  100 - 108 , etc. Printhead  108  is shown being maintained by maintenance element  134  in FIG. 4. Element  134  is shown as fluid cleaning system for printhead  108  including fluid source  136 . In this station, the printhead can be serviced through various means such as the ability to cap the printhead with an appropriate capping means that will prevent the printheads from drying out during non-printing cycles. This station may also contain appropriate means such as controlled vacuum  150  linked to recess  152  to prime the printhead with bio-specific fluids (shown in FIG. 6) or the ability to jet into an element  134  that contains an absorbing material  160  as shown in FIG. 7.  
         [0057]    In addition, a maintenance element  134  is shown in FIG. 8 wherein said element contains a flexible wiping member  108  in which the motion of the printhead  108  relative to said wiping member  170  maintains the surface of the printhead  108  free of fluids and dirt.  
         [0058]    The printheads  100 - 108  (or support) will move relative to each other to create the required deposition pattern, which could include but not limited to a uniform distribution of bio-specific sites, or groupings  202  of bio-specific sites that might repeat across the surface defined by the support. FIG. 9 shows a pattern in which groupings or sub-regions of the array  200  are arranged into a pattern of sub-arrays. This sub-array pattern is preferably contains 10 unique bio-specific fluids, more preferably contains 100 unique bio-specific fluids, and most preferably contains 1000 unique bio-specific fluids.  
         [0059]    As shown in FIG. 10, the spacing of the bio-specific fluid spots  206  in the sub-array  204  preferably has a dot spacing (dXs, dYs) of 3000 um, more preferably of 1000 um, and most preferably of 300 um. Additionally, as shown in FIG. 1, the groupings of bio-specific sites (sub-arrays)  202  are most preferably arranged with a spacing (dX, dY) of 1 cm.  
         [0060]    The printheads in the system can produce droplet sizes that are commensurate with the dot sizes as mentioned above. Specifically, as shown in FIG. 12, a bio-specific fluid droplet  402  of appropriate volume shall be produced by printhead  300  to create the sub-array. The volume of the droplet can be determined by first characterizing the fluid spread on the receiving layer as defined by the Spread Factor,  
         Spread Factor= SF =Dot  dia /Drop  dia    
         [0061]    Once this has been determined, then the appropriate drop volume can be calculated (assuming a spherical drop relationship),  
           Vol=( 4π/3)* R   3 =(4π/3)*((Dot  Dia )/(2 SF))   3    
         [0062]    where R is the radius of the drop.  
         [0063]    Assuming a Spread Factor of 2, then preferably, this volume to produce the sub-array shall be 200 nL, or more preferably 7.5 nL, and most preferably 200 pL micro-droplets.  
         [0064]    [0064]FIG. 13 shows a printhead  400  for large droplets  402  that can cover the entire sub-array  404  with a single fluid.  
         [0065]    The device can have printheads that can produce micro droplet volumes that are appropriate for generating sub-arrays as well as ones that can produce macro-droplet volumes for covering the sub-array fluids with yet another fluid, which will increase the bio-diversity of the array. In practice, it is envisioned that an array is initially created, with 1000 unique bio-specific fluids in an N×M pattern. This is defined as a sub-array as shown in FIG. 9. The printhead that generates this sub-array is capable of generating micro-droplets. It is further envisioned that another printhead, capable of producing macro-droplets, will further increase the bio-diversity or search capabilities of the array by placing a droplet that covers the entire sub-array region, as thus interacts with the entire unique bio-fluids contained in this N×M sub-array.  
         [0066]    The invention has been described in detail with particular reference to certain preferred embodiments thereof, but it will be understood that variations and modifications can be effected within the spirit and scope of the invention.  
       Parts List  
       [0067]    block of ferroelectric material  
         [0068]    [0068] 10  cylindrical void  
         [0069]    [0069] 12  electrodes  
         [0070]    [0070] 20 , 22  glass or plastic capillary  
         [0071]    [0071] 30  flexible or rigid tubing  
         [0072]    [0072] 32  orifice plate  
         [0073]    [0073] 40  orifices  
         [0074]    [0074] 42  piezelectric element  
         [0075]    [0075] 50  capillary elements  
         [0076]    [0076] 52  piezelectric element  
         [0077]    [0077] 60  capillary elements  
         [0078]    [0078] 62  ferroelectric material  
         [0079]    [0079] 70  cylindrical voids  
         [0080]    [0080] 74  orifice plate  
         [0081]    [0081] 76  apertures  
         [0082]    [0082] 90  printing station  
         [0083]    [0083] 100 - 108  page-width printhead  
         [0084]    [0084] 110  fluid delivery system  
         [0085]    [0085] 111  computer  
         [0086]    [0086] 112  fluid delivery system  
         [0087]    [0087] 114  service station  
         [0088]    [0088] 116 - 118  controls mechanism  
         [0089]    [0089] 120  printhead direction  
         [0090]    [0090] 122  printhead direction  
         [0091]    [0091] 130  nozzles  
         [0092]    [0092] 134  fluid cleaning system  
         [0093]    [0093] 136  fluid source  
         [0094]    [0094] 150  controlled vacuum  
         [0095]    [0095] 152  recess  
         [0096]    [0096] 160  absorbing material  
         [0097]    [0097] 170  wiping member  
         [0098]    [0098] 200  array  
         [0099]    [0099] 202  groups of bio-specific sites  
         [0100]    [0100] 204  sub-array  
         [0101]    [0101] 206  bio-specific fluid spots  
         [0102]    [0102] 300  printhead  
         [0103]    [0103] 402  bio-specific fluid droplet  
         [0104]    [0104] 404  entire sub-array