Patent Publication Number: US-2005136546-A1

Title: Microtiter plate, system and method for processing samples

Description:
BACKGROUND OF THE INVENTION  
      Microtiter plates are multi-well plates that are adapted for receiving samples to be processed at a plurality of wells. Each well defines a reaction site where a sample is usually mixed with one or more reagents in order to form a sample-reagent mixture which is the subject to analysis e.g. by means of a photometer or a fluorometer.  
      In recent developments in the field of processing large numbers of samples that have a liquid component or a liquid and a solid component or a liquid and a gel component there is a need for a device that makes possible to separate the liquid from the solid or gel component of each sample rapidly and at low cost. There is in particular a need for a device of this kind which is suitable for processing in the latter way individual samples of very low volume, e.g. lower than 30 microliter.  
     SUMMARY OF THE INVENTION  
      The invention provides a microtiter plate that is configured and dimensioned for performing the above-mentioned separations for a large number of samples rapidly and at low cost.  
      The invention also concerns a microtiter plate for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component.  
      The invention further concerns a system for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component.  
      The invention further concerns a method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component.  
      According to a first aspect of the invention the above aim of the invention is attained with a microtiter plate of the above mentioned kind comprising 
          a single piece body, which is made by injection molding     said body having an array of cavities and 
            each of said cavities having an open upper end and a closed bottom end,     each of said cavities having a bottom inner surface and comprising a first chamber for receiving a predetermined volume of a sample to be processed, a second chamber and a passage which fluidically connects said first and second chambers with each other, said passage having a top opening,     said first chamber, said second chamber and said passage having each a bottom inner surface which is portion of the bottom inner surface of said cavity,     a region in the lower part of said passage being adjacent to the bottom end of the cavity, said region being so configured and dimensioned that it allows passage of liquid from one of said chambers to the other only when a centrifugal force is applied to the microtiter plate, but does not allow passage of any solid or gel component the size of which is larger than the width of said region.    
               

      According to a second aspect of the invention the above aim of the invention is attained with a system for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, said system comprising a microtiter plate according to the invention.  
      According to a third aspect of the invention the above aim of the invention is attained with a method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, said method comprising 
          (a) introducing a predetermined volume of a sample, for example a sample comprising one or more biomolecules, having a liquid component or a liquid and a solid component or a liquid and a gel component into a first chamber of a cavity of a microtiter plate according to the invention,     (b) centrifugating the microtiter plate for transferring liquid from said first chamber to said second chamber, the liquid component of said sample being thereby entirely removed from said first chamber leaving therein only the solid or gel component of the sample.        

      According to a fourth aspect of the invention the above aim of the invention is attained with a method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, said method comprising 
          (a) introducing a predetermined volume of a sample, for example a sample comprising one or more biomolecules, having a liquid component or a liquid and a solid component or a liquid and a gel component into a first chamber of a cavity of a microtiter plate according to the invention,     (b) fluidically connecting one end of a pipetting tip with a second chamber of a cavity of a microtiter plate according to the invention,     (c) performing pipetting operations on said sample with said pipetting tip for either transferring liquid from said first chamber to said second chamber or for adding and/or removing a liquid to respectively from said first chamber and/or said second chamber.       

    
    
     BRIEF DESCRIPTION OF THE FIGURES  
      The subject invention will now be described in terms of its preferred embodiments with reference to the accompanying drawings. These embodiments are set forth to aid the understanding of the invention, but are not to be construed as limiting.  
       FIG. 1  shows a perspective view of a microtiter plate  11  according to the invention.  
       FIG. 2  shows an enlarged view of part  11  of microtiter plate  11  in  FIG. 1 .  
       FIG. 3  shows a partial cross-sectional view of microtiter plate  11  along plane III-III in  FIG. 2 .  
       FIG. 4  shows the same view of microtiter plate  11  as  FIG. 3   a , but shows in addition a pipetting tip inserted in chamber  17 .  
       FIG. 5  shows a partial cross-sectional view of microtiter plate  11  along plane V-V in  FIG. 2 .  
       FIG. 6  shows an enlarged cross-sectional view of a part of  FIG. 3 .  
       FIG. 7  shows a partial cross-sectional view of microtiter plate  11  along plane VI-VI in  FIG. 6 .  
       FIG. 8  shows a partial cross-sectional view of microtiter plate  11  along plane VII-VII in  FIG. 6 .  
       FIG. 9  shows a top view of a portion of microtiter plate  11  in  FIG. 1 . 
    
    
     REFERENCE NUMBER LIST  
     
         
          11 microtiter plate  
          12 single piece body  
          13 cavity  
          14 upper end of cavity  13   
          15 bottom end of cavity  13   
          16 first chamber of cavity  13   
          17 second chamber of cavity  13   
          18 passage  
          19 top opening of passage  18   
          21 zone of passage  18   
          22 bottom of second chamber  17   
          23 bottom of first chamber  16   
          24 top side of microtiter plate  11   
          25 coating of bottom of passage  18   
          26 zone of minimum width of passage  18   
          27 circular line portion  
          28 circular line portion  
          29 bottom of passage  18   
          31 curved line portion  
          32 curved line portion  
          33 pipetting tip  
          34 sealing means  
          35 side edge of microtiter plate  11   
          36 side edge of microtiter plate  11   
          37 solid element  
       
    
     DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS  
     DETAILED DESCRIPTION  
      The invention concerns a microtiter plate for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, a system for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, and a method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component.  
      According to a first aspect of the invention the above aim of the invention is attained with a microtiter plate of the above mentioned kind comprising 
          a single piece body, which is made by injection molding,     said body having an array of cavities and 
            each of said cavities having an open upper end and a closed bottom end,     each of said cavities having a bottom inner surface and comprising a first chamber for receiving a predetermined volume of a sample to be processed, a second chamber and a passage which fluidically connects said first and second chambers with each other, said passage having a top opening,     said first chamber, said second chamber and said passage having each a bottom inner surface which is portion of the bottom inner surface of said cavity,     a region in the lower part of said passage being adjacent to the bottom end of the cavity, said region being so configured and dimensioned that it allows passage of liquid from one of said chambers to the other only when a centrifugal force is applied to the microtiter plate, but does not allow passage of any solid or gel component the size of which is larger than the width of said region.    
               

      According to a second aspect of the invention the above aim of the invention is attained with a system for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, said system comprising a microtiter plate according to the invention.  
      According to a third aspect of the invention the above aim of the invention is attained with a method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, said method comprising 
          (a) introducing a predetermined volume of a sample, for example a sample comprising of one or more biomolecules, having a liquid component or a liquid and a solid component or a liquid and a gel component into a first chamber of a cavity of a microtiter plate according to the invention,     (b) centrifugating the microtiter plate for transferring liquid from said first chamber to said second chamber, the liquid component of said sample being thereby entirely removed from said first chamber leaving therein only the solid or gel component of the sample.        

      According to a fourth aspect of the invention the above aim of the invention is attained with a method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component, said method comprising 
          (a) introducing a predetermined volume of a sample, for example a sample comprising of one or more biomolecules, having a liquid component or a liquid and a solid component or a liquid and a gel component into a first chamber of a cavity of a microtiter plate according to the invention,     (b) fluidically connecting one end of a pipetting tip with a second chamber of a cavity of a microtiter plate according to the invention,     (c) performing pipetting operations on said sample with said pipetting tip for either transferring liquid from said first chamber to said second chamber or for adding and/or removing a liquid to respectively from said first chamber and/or said second chamber.        

      The following examples are provided for illustrative purposes and are not intended to limit the scope of applicants&#39; invention.  
     EXAMPLE OF A MICROTITER PLATE FOR PROCESSING SAMPLES  
       FIG. 1  shows a microtiter plate  11  according to the invention for processing samples comprising biomolecules to be analyzed, and further having a liquid component or a liquid and a solid component or a liquid and a gel component.  
      A sample of the above mentioned kind is processed in a cavity of the microtiter plate and such processing includes steps which have the effect of extracting biomolecules to be analyzed from the solid or gel component of the sample and transferring those biomolecules to the liquid component of the sample. After this step the liquid component carrying the biomolecules to be analyzed is separated from the solid or gel component of the sample as described in detail hereinafter. This separation is the main aim of the instant invention.  
      The term “liquid component” comprises any liquid containing biomolecules in solution or any liquid containing a solid component and/or a gel component to form a suspension.  
      The term “solid component” comprises a solid in the suspension that shall be separated by the proposed structure and method, e.g., chromatography beads.  
      The term “gel component” comprises one or more pieces of agarose or polyacrylamide gel or another gel. The size of the gel component is limited by the size of the cavity and the size of the connecting structure between the two cavities.  
      The maximum size of the solid or gel component of the sample is limited by the size of the cavity of the microtiter plate. The minimum size of the solid or gel component that can be separated from the liquid component is limited by the size of the passage which connects said first and second chamber of the cavity with each other.  
      The term “biomolecule” comprises all organic molecules, including macromolecules, found in living organisms and in particular, proteins, peptides, DNA, RNA and metabolites thereof.  
      As used herein, the term “having” is equivalent to the term, and shall have the meaning of the term, comprising.  
      Microtiter plate  11  comprises a single piece body  12 . The single piece body  12  is made by injection molding of a suitable plastic material, e.g. Polypropylene (PP), Cyclic Olefin Copolymer (COC), Acrylonitrile/Butadien/Styrene (ABS), Polycarbonate (CC) or Polystyrene (PS), or of other materials known to one of ordinary skill in the art.  
      Body  12  has an array of cavities  13  and side edges  35 ,  36 . In a preferred embodiment, the grid spacing is of e.g. 4.5 millimeter measured along each of edges  35 ,  36 , i.e. in both X-direction and Y-direction shown by arrows in  FIGS. 1 and 9 .  
      As shown in particular by  FIGS. 1 and 9 , the cross-section of each of cavities  13  has a length axis which forms an angle A of about 45 degrees with a side edge  35 ,  36  of the microtiter plate  11 . This spatial arrangement of cavities makes possible to form a relatively large number of such cavities in a microtiter plate of standard size as known to one of ordinary skill in the art. In a preferred embodiment, the standard size plate has e.g. a length of about 127.76±0.25 millimeter and a width of about 85.48±0.25 millimeter.  
      In a preferred embodiment, single piece body  12  has standard outer dimensions of a microtiter plate and comprises 384 cavities  13 . In another preferred embodiment, single piece body  12  has standard outer dimensions of a microtiter plate and comprises 1536 cavities  13 . In yet another preferred embodiment, single piece body  12  has standard outer dimensions of a microtiter plate and comprises 96 cavities.  
      As shown in particular by  FIGS. 1, 2  and  9  each of cavities  13  has an inner surface the cross-section of which is a closed curve and the inner surface has no corner or sharp edge. In a preferred embodiment the closed curve has approximately the shape of two circular line portions  27 ,  28  connected with each other by curved line portions  31 ,  32 .  
      As shown by FIGS.  3  to  8 , each of cavities  13  has an open upper end  14  and a closed bottom end  15  and each of cavities  13  has a bottom inner surface and comprises a first chamber  16  for receiving a predetermined volume of a sample to be processed, a second chamber  17  and a passage  18  which fluidically connects chambers  16  and  17  with each other. Passage  18  has a top opening  19 . The total volume of a cavity  13  is e.g. about 30 microliter. The bottom  23  of chamber  16 , the bottom  22  of chamber  17  and the bottom of passage  18  have each an inner surface which is a portion of the inner surface of the bottom  15  of cavity  13 .  
      Chambers  16 ,  17  and passage  18  have side walls with an inclination angle of about 4 degrees.  
      In a preferred embodiment, chamber  16  is adapted for receiving a sample having a liquid component or a liquid and a solid component or a liquid and a gel component, whereas chamber  17  is adapted for receiving a pipetting tip  33  shown by  FIG. 4 .  
      In a preferred embodiment microtiter plate  11  further comprises sealing means  34 , shown in  FIG. 4 , which seal the contact surface of tip  33  with the microtiter plate  11  and second sealing means (not shown) which seal the top opening of passage  18 .  
      As shown in particular by  FIGS. 1, 2  and  9 , passage  18  has a variable width in a direction extending from chamber  16  to chamber  17  and that width has a minimum at a zone  26  located between chambers  16  and  17 .  
      A region  21  in the lower part of passage  18  is adjacent to the bottom end  15  of the cavity  13 . Region  21  is so configured and dimensioned that it allows passage of liquid from one of chambers to the other only when a centrifugal force is applied to the microtiter plate, but does not allow passage of any solid or gel component the size of which is larger than the width of region  21 .  
      In a preferred embodiment, region  21  of passage  18  is configured and dimensioned as a capillary passage adapted for supporting or facilitating flow of liquid from one of chambers  16 ,  17  to the other. This is for instance the case when the entire length of region  21  is a capillary adapted for receiving liquid and is thereby able to provide a fluidic connection between the bottom of chamber  16  and the bottom of chamber  17 . The bottom of passage  18  (shown in  FIG. 7 ) has a radius R 1 , e.g. R 1 = 0 . 3  millimeter. The radius R 1  is preferably comprised e.g. in a range between 0.1 to 0.5 millimeter.  
      In another preferred embodiment, region  21  of passage  18  is configured and dimensioned as a capillary passage adapted for preventing a displacement of a solid or gel component of the sample through passage  18 .  
      As shown by FIGS.  3  to  6 , in a preferred embodiment the bottom  22  of chamber  17  lies at a lower level than the bottom  23  of first chamber  16  when the microtiter plate  11  is in horizontal position and the upper ends  14  of chambers are on the top side  24  of the microtiter plate  11 . As shown by  FIG. 6 , the bottom of chamber  16  has an inclination of about 20 degrees with respect to the top side  24  of plate  11 . As shown by  FIG. 7 , the deepest point of the bottom of chamber  17  has a depth H 1 . As shown by  FIG. 8 , chamber  16  has a depth H 2 , that is smaller than that of H 1  (for example, H 1 =about 5 mm, H 2 =about 4 mm). The depth of H 1  and H 2 , respectively are determined by the taper of the body chambers, the distance from chamber  16  to  17 , and the angle of inclination in chamber  17 .  
      In a preferred embodiment the inner surface of the bottom  29  of passage  18  which fluidically connects chambers  16  and  17  with each other has a shape that contributes to maximize the centrifugal force exerted on a sample contained in first chamber  16  when microtiter plate  11  is centrifuged by means of a centrifugation apparatus.  FIG. 6  shows such a shape of the bottom  29  of passage  18 .  
      In a preferred embodiment of microtiter plate  11  at least a portion of the inner surface of the bottom of each of said cavities  13  is a hydrophilic or hydrophobic surface, or is a surface having a hydrophilic or hydrophobic coating. The purpose of these surface properties is to create flow conditions that are suitable for the intended use of the microtiter plate, e.g. when a preferred sense of flow is suitable for the desired liquid handling process.  
      In a preferred embodiment at least a portion of or the entire inner surface of the bottom  29  of passage  18  is a hydrophilic surface or is a surface having a hydrophilic coating  25  shown by  FIG. 6 . This feature facilitates the flow of liquid through passage  18  and thereby ensures that the entire volume of liquid in chamber  16  is transferable to chamber  17  by centrifugation of microtiter plate  11 .  
      In a preferred embodiment at least a portion of or the entire inner surface of the bottom  23  of chamber  16  is a hydrophilic surface or is a surface having a hydrophilic coating (not shown). This feature facilitates the flow of liquid from chamber  16  to passage  18  and thereby ensures that the entire volume of liquid in chamber  16  is transferable to chamber  17  by centrifugation of microtiter plate  11 .  
      In a preferred embodiment at least a portion of or the entire inner surface of the bottom  22  of chamber  17  is a hydrophobic surface or is a surface having a hydrophobic coating (not shown). This feature facilitates the flow of liquid from chamber  16  to passage  18  and thereby ensures that the entire volume of liquid in chamber  16  is transferable to chamber  17  by centrifugation of microtiter plate  11 .  
      As shown by FIGS.  3  to  8 , in a preferred embodiment each of cavities  13  tapers towards its bottom end  15 , i.e. the cross-section of each cavity  13  diminishes towards the bottom thereof.  
      As shown by  FIG. 9 , in a preferred embodiment a solid element  37 , which is liquid permeable, is arranged in region  21  of passage  18 .  
      Solid element  37  is e.g. a filter element having a porous structure that allows passage of particles having a size that is smaller than a predetermined size. Such a filter element is made e.g. of glass or of a plastic material or of other similar materials. In a preferred embodiment, solid element  37  is a membrane that allows passage of particles having a size that is smaller than a predetermined size. Such membrane is made e.g. of a plastic material, paper, a gel or a microfiber or of other materials as known to one of ordinary skill in the art.  
      In a preferred embodiment solid element  37  is a test element, e.g. a chromatographic test element. Test element  37  is e.g. a membrane or a strip similar to a chromatographic strip which in a first step is able to retain a sample material of a certain kind as a sample flows from chamber  16  to chamber  17  through passage  18  and in a subsequent step is able to release that sample material when said test element is brought in contact with a suitable reagent, the released sample and reagent mixture being then transferable to chamber  17  e.g. by centrifugation of plate  11 .  
      In a preferred embodiment solid test element  37  or at least a portion thereof is a coating having hydrophilic properties or hydrophobic properties. The coating is selected based upon the properties of the sample, such as a biomolecule, to be processed, so that the sample, e.g., one or more biomolecules, preferably bind to the coating and/or otherwise do not pass through solid element  37 , e.g., the filter.  
      In a preferred embodiment, solid element  37  is a filter or test element that at least a portion thereof has a coating having hydrophilic or hydrophobil properties.  
     EXAMPLE 1 OF A SYSTEM FOR SAMPLE PROCESSING  
      According to the invention a first system for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component comprises a microtiter plate  11  of the kind described above with reference to  FIGS. 1-9 .  
      In a preferred embodiment this first system further comprises a centrifugation apparatus (not shown in the drawings) for centrifugating the microtiter plate  11 .  
     EXAMPLE 2 OF A SYSTEM FOR SAMPLE PROCESSING  
      According to the invention a second system for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component comprises a microtiter plate  11  of the kind described above with reference to  FIGS. 1-8 .  
      In a preferred embodiment this second system further comprises a pipetting tip  33  (shown in  FIG. 4 ) which is insertable into chamber  17  and which is connectable to a pipetting apparatus including overpressure or underpressure generating means.  
     EXAMPLE 1 OF A METHOD FOR SAMPLE PROCESSING  
      According to the invention a first method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component comprises 
          (a) introducing a predetermined volume of a sample having a liquid component or a liquid and a solid component or a liquid and a gel component into chamber  16  of a cavity  13  of a microtiter plate  11  of the above-described type,     (b) centrifugating the microtiter plate  11  for transferring liquid from chamber  16  to chamber  17 , the liquid component of sample being thereby entirely removed from first chamber  16  leaving therein only the solid or gel component of the sample.        

      In a preferred embodiment, the above-mentioned transfer of liquid is effected exclusively by means of centrifugal force generated by centrifugation of the microtiter plate  11 . The sample volume transferred from chamber  16  to chamber  17  by centrifugation is in the range of about e.g. 0.05 to 2 microliter.  
     EXAMPLE 2 OF A METHOD FOR SAMPLE PROCESSING  
      According to the invention a second method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component comprises 
          (a) introducing a predetermined volume of a sample having a liquid component or a liquid and a solid component or a liquid and a gel component into a first chamber  16  of a cavity  13  of a microtiter plate  11  of the above-described type,     (b) fluidically connecting one end of a pipetting tip  33  with a second chamber  17  of a cavity  13  of a microtiter plate  11  of the above-described type,     (c) connecting another end of pipetting tip  33  with a pipetting apparatus including underpressure generating means for aspirating and thereby removing the liquid component of said sample from said first chamber  16  and leaving therein only the solid or gel component of the sample.        

     EXAMPLE 3 OF A METHOD FOR SAMPLE PROCESSING  
      According to the invention a third method for processing samples having a liquid component or a liquid and a solid component or a liquid and a gel component comprises 
          (a) introducing a predetermined volume of a sample having a liquid component or a liquid and a solid component or a liquid and a gel component into a first chamber  16  of a cavity  13  of a microtiter plate  11  of the above-described type,     (b) fluidically connecting one end of a pipetting tip  33  with a second chamber  17  of a cavity  13  of a microtiter plate  11  of the above-described type,     (c) performing pipetting operations on said sample with pipetting tip  13  for either transferring liquid from first chamber  16  to second chamber  17  or for adding and/or removing a liquid to respectively from said first chamber  16  and/or said second chamber  17 .        

     EXAMPLE OF USE OF THE MICROTITER PLATE, SYSTEM AND METHOD ACCORDING TO THE INVENTION  
      In a preferred use of the microtiter plate, system and method according to the invention the gel component of the sample contains biomolecules to be analyzed.  
      Proper use of the microtiter plate according to the invention is subject to the condition that the volume of sample introduced into chamber  16  is smaller than a predetermined maximum value. When this condition is fulfilled only the liquid component of the sample passes through region  21  of passage when transferred from chamber  16  to chamber  17  and any solid or gel component of the sample remains in chamber  16 . If the above mentioned condition is not fulfilled, some of the solid and/or gel components of the sample can pass from chamber  16  to chamber  17  through the upper part of passage  18  and the desired separation of the liquid from the solid and/or gel components of the sample is not or not completely achieved. Therefore, in the above described methods the predetermined volume of the sample introduced into chamber  16  is smaller than a predetermined maximum value determined by the shape and the dimensions of the cavity  13 , the chambers  16  and  17 , and the passage  18 .  
      Although preferred embodiments of the invention have been described using specific terms, such description is for illustrative purposes only, and it is to be understood that changes and variations may be made without departing from the spirit or scope of the following claims.