Abstract:
A multi-well assay plate structure ( 54 ) and assay apparatus and a method for performing chemical biochemical assays is described. The multi-well assay plate structure ( 54 ) defines a relatively shallow substantially enclosed space ( 71 ) above a plurality of wells ( 76 ), with the enclosed space ( 71 ) having an inlet ( 72 ) and an outlet ( 22 ) separate from the inlet. Fluid introduced via the inlet ( 72 ) flows into the space ( 71 ) and/or wells ( 76 ) by displacing air. Withdrawal of the fluid via the inlet ( 72 ) or outlet leaves fluid in the wells ( 76 ) allowing various tests to be performed. Various embodiments of the structure are described. The preferred arrangement embodies the structure on a transparent plastic disk which can be used with automatic fluid handling apparatus ( 80 ) and the results assessed using optical assessment apparatus ( 81 ). The apparatus can be used to perform a variety of assays but, in particular, biochemical/chemical assay, immunoassays and genetic (DNA) assays and it can be used in a laboratory for multiple sample testing or at a point-of-care, i.e. in a surgery or clinic.

Description:
BACKGROUND OF THE INVENTION 
       [0001]    1. Field of the Invention 
         [0002]    The present invention relates to apparatus and to a method for conducting assays and, in particular, to multi-well plate structures for receiving and holding, in separate wells, volumes of liquid for the purpose of conducting chemical or biochemical assays. Multi-well trays or plates having a 2-dimensional array of small wells are commonly used in medicine and science to facilitate testing of a liquid analyte. One particular area of use is blood screening where blood or blood products are introduced into the wells to test for viruses such as HIV, hepatitis etc. 
         [0003]    2. Description of the Related Art 
         [0004]    Such tests (immunoassays) typically involve an antigen-antibody interaction, where the surfaces of the wells are coated with specific antigen itself. This approach detects circulating antibodies to that specific antigen. Alternatively the wells can be coated with a specific antibody which captures circulating antigen which is, in turn, identified by a second antibody directed against a second epitope on the captured antigen. These two approaches are just two of the large number of variants developed in immunoassay (review Principles and Practice of Immunoassay Price &amp; Newman 1997 ISBN 1-56159-145-0). 
         [0005]    In an immunoassays sample must be applied and in most cases subsequent addition of reagents or washing buffer is required. Typically the well is exposed to blood or blood product and the well is rinsed clean and a further reactant, which binds either to exposed antibodies or captured antigens is introduced into the wells, to create an observable reaction. These reactions may produce a color or some other observable change. This enables the wells containing specific antigen antibody reactions to be identified and the extent of these reactions quantified. 
         [0006]    It is often necessary to fill each well of a multi-well tray with a precisely defined volume of analyte. This is normally achieved using a single or multi-headed micro-pipette. However, this process is often time consuming and, particularly where a large number of wells are to be filled, can lead to a number of wells being missed. 
         [0007]    In certain circumstances it is necessary that the wells of a tray be contained within a substantially closed container, e.g. to avoid the risk of contamination of the wells and of leakage of contaminated material. With trays such as this, it may be difficult or impossible to gain access to the wells to enable them to be filled using a micro-pipette. 
       SUMMARY OF THE INVENTION 
       [0008]    It is an object of the present invention to overcome or at least mitigate the disadvantages of known multi-well trays. 
         [0009]    This is achieved by providing a multi-well assay plate structure which defines a relatively shallow substantially enclosed space above a plurality of wells, with the enclosed space having an inlet and an outlet separate from the inlet. Fluid introduced via the inlet flows into the space, and covers the wells, by displacing air. Withdrawal of the fluid from the space via the inlet or outlet leaves fluid in the wells allowing various tests to be performed. 
         [0010]    According to a first aspect of the present invention there is defined a multi-well assay plate structure comprising:
       a first upper surface,   a second lower surface having a plurality of wells disposed therein,   the first and second surfaces defining a chamber having an inlet and an outlet, the inlet and outlet allowing fluid to be introduced and withdrawn from the chamber, the wells being proportioned and dimensioned to retain a volume of fluid in each well following withdrawal of the liquid.       
 
         [0014]    Preferably, the chamber is shallow enough to allow fluid to fill the wells and the chamber. The wells are deep enough to retain a volume of fluid following withdrawal of fluid in the space above the wells. 
         [0015]    The plate structure can be of any convenient shape but, advantageously, is sector-shaped with a detachable handle at the longer arc-portion to facilitate locating the sector on a disc. Conveniently, a plurality of sector-shaped structures are located on the disc. 
         [0016]    Conveniently, also the sectors and discs are made of plastic and the sectors can be snap-fitted onto the disc. The sectors and the disc include lock and key portions to allow the sectors to be snap-fitted in the correct orientation only. 
         [0017]    Alternatively, a disc with a plurality of separate sections can be manufactured or molded in one piece instead of snap-in sectors. 
         [0018]    The composite structure may be snap-fitted onto a compact disk. 
         [0019]    The disk structure may have a circumferential gutter extending around its periphery to facilitate collection of fluid following fluid introduction/withdrawal from the chamber. 
         [0020]    The wells are dimensioned and proportioned in terms of diameter and depth to receive and retain fluid containing the analyte or part of the reagent under test. The exact dimensions are a matter of choice and depend on a number of parameters such as the type of material of the surfaces of the chamber and wells; viscosity of the fluid and the depth (height) of the space between the first and second surfaces. 
         [0021]    Advantageously, the dimensions of the structure are such that the wells fill to retain sufficient fluid the space is flooded and withdrawal to allow a measurable reaction to be measured within an individual well without contribution from adjacent wells. The overall process of sequential steps of flood and fill is advantageous in that it allows both discrete measurements within individual wells when filled and efficient washing of an array of wells (flood) which is useful in multistep procedures, such as immunoassays, which requires sequential application of reagents interspersed with rigorous washing steps. This permits the wells to be cleaned or rinsed in the same way as filling to allow subsequent tests to be carried out within an individual well whilst avoiding cross-contamination between adjacent wells. 
         [0022]    The structure is preferably made of transparent or otherwise optically transmissive plastic to facilitate optical reading of the wells to determine the results of the tests. Conveniently, the structure is integrated with automatic fluid handling apparatus and an optical reader to allow automatic fluid handling and optical assessment of the results of the reactions. Alternatively, fluid handling can be manually controlled and the results of the reactions within the structure can be assessed by an optical reader or be scored by visual assessment. 
         [0023]    According to a second aspect of the present invention there is provided a multi-well assay structure comprising an upper surface and a lower closely spaced opposed surface, said upper and lower surfaces defining a relatively shallow space therebetween, the lower surface having a plurality of wells therein, at least two spaced apart openings providing access to said space from an external location, wherein a fluid introduced into said space through one of said openings fills substantially all of the space and covers of the wells and said fluid, when subsequently withdrawn through the same or the other opening, leaves the wells filled with liquid. 
         [0024]    The volume of fluid introduced into each well when using the structure of the present invention is substantially defined by the volume of the well. The accuracy and precision with which the wells can be filled is therefore defined by the accuracy and precision with which the wells can be fabricated and which is generally high. Furthermore, the multiplicity of wells can be filled by way of a single injection and withdrawal of fluid through an opening into the space containing the wells, so that the wells can be filled extremely rapidly. 
         [0025]    The structure of the present invention provides for the filling of a plurality of wells m a substantially closed chamber, the only openings into that container being the fluid injection opening and a second ‘vent’ opening. 
         [0026]    The structure of the present invention simplifies the process of cleaning or rinsing previously filled wells as this can be achieved by repeatedly injecting and withdrawing fluid through one of said openings. 
         [0027]    Conveniently, the spacing between said upper and lower surfaces is sufficiently small to facilitate the flow of fluid m said space by capillary or capillary like action. Typically, the spacing is less than 1 mm and preferably less than 0.5 mm. 
         [0028]    Preferably, said upper and lower surfaces are substantially planer. 
         [0029]    The wells may have any suitable geometry. For example, the wells may be provided m said lower surface by blind circular holes with a semi-spherical termination. Alternatively, the wells may have substantially straight sidewalls, e.g. so that the sidewalls extend substantially vertically and terminate m a flat base Vertical sidewalls assist m preventing the transfer of fluid between adjacent wells. 
         [0030]    The surfaces may be provided by respective upper and lower plates which are spaced apart by one or more spacer walls. 
         [0031]    Preferably, the opening through which fluid is introduced into said space is provided through either the upper or lower surface and, more preferably, through the upper surface. The additional opening may be provided through said upper or lower surface or through a side surface. 
         [0032]    Preferably, said opening for introducing a fluid comprises a relatively small opening arranged to receive the end of a syringe or similar liquid injecting device, where the opening forms a substantially air-tight seal around said end. 
         [0033]    Preferably, said lower surface of the container is treated to increase the hydrophobicity to facilitate smooth flow of liquid across the sector and hydrophilicity to aid movement of liquid into desired locations, e.g. wells. This helps to, prevent the formation of air pockets in the space and aids filling of the wells. The treatment may comprise for example exposing the surface to a wetting agent, e.g. poly-1-lysine, or exposing the surface to a gas plasma. 
         [0034]    In one embodiment of the present invention, the multi-well structure is embodied in a disc. The disc effectively comprises upper and lower circular plates, the internal surfaces of which respectively define said upper and lower opposed surfaces. Preferably, said opening for introducing liquid into the space is a hole passing through the upper circular plate. Preferably, the second opening is provided at the peripheral edge of the disc. The space between the upper and lower plates is subdivided, by one or more dividing walls, to provide a plurality of multi-well plates in which case each space is provided with an opening and a vent to enable each space to be independently filled. The dividing walls may extend radially and/or may be concentric to one another. 
         [0035]    Preferably, at least one of the upper and lower plates forming the container are transparent to enable optical inspection of the wells from outside the container. The other of the upper and lower plates may comprise a reflecting surface so that radiation entering into the container through the transparent plate transverses the container in both directions, resulting in an improved signal detection for optical inspection. 
         [0036]    In an alterative embodiment of the present invention there is provided a disc arranged to receive a plurality of sector (pie) shaped inserts each of which comprises a generally planar upper surface having a plurality of wells provided therein. For each insert, the disc comprises a substantially planar surface arranged, in use, to oppose said substantially planar insert surface and means for retaining the insert in position so that the respective planar surfaces are in closely spaced opposition to one another, and said at least two openings. 
         [0037]    Preferably, the opening for filling the container is provided through the planar surface of the disc. The vent opening is preferably provided at, or adjacent to, the peripheral edge of the disc. 
         [0038]    The disc preferably comprises upper and lower circular plates separated by radially extending spacers. The spacers define slots between the plates for receiving said inserts. Preferably, said planar surface of each insert comprises upstanding walls around at least a portion of its periphery for the purpose of sealing the inner edges of the insert to the opposed planar surface of the disc, thereby to prevent seepage of liquid around the insert. 
         [0039]    According to a third aspect of the present invention there is provided a method of filling the wells of the multi-well structure of the above first aspect of the present invention, said method comprising the steps of:
       introducing a fluid into said chamber through one of said openings to substantially flood the chamber; and   subsequently withdrawing the fluid from the chamber through the same or the other opening to leave liquid in the wells.       
 
         [0042]    Preferably, the method further includes the step of forming an air tight seal between the fluid inlet and an end region of a syringe or similar liquid injecting device, and injecting fluid through the opening into the chamber and subsequently sucking liquid out of the space through the opening. 
         [0043]    According to a fourth aspect of the present invention there is provided a method of conducting a chemical or biochemical assay said method comprising the steps of:
       providing a surface within a substantially enclosed chamber having a plurality of wells at spaced locations sufficient to allow a reaction at each well location,   treating each well with a first reagent, flooding the enclosed chamber and covering the wells with a fluid carrying at least a second reagent,   removing excess fluid from said chamber to leave a mixture of said first and second reagents in each well, and   optically assessing each well and determining if a reaction occurred and correlating the reaction results to provide an assay of the chemical or biochemical reactions under test.       
 
         [0048]    Preferably, the step of optical assessment is carried out automatically using optical reading apparatus. 
         [0049]    Preferably also, the surfaces with the wells having first fluid carrying reagents are prior prepared for loading into the structure. 
         [0050]    Conveniently, the fluid carrying at least the second reagent is introduced into the structure and withdrawn from the structure using suitable automatic fluid handling apparatus. 
         [0051]    Conveniently also, after optical assessment of the results of the assay, the automated fluid handling apparatus is used to inject and withdraw rinsing fluid a predetermined number of times from the well tray to clean the wells for receiving subsequent samples for assay. 
         [0052]    According to a fifth aspect of the present invention, there is provided chemical/biochemical assay apparatus comprising
       an assay plate structure defined in said first aspect and having a plurality of wells for receiving samples to be assayed,   fluid handling means for introducing and removing fluid reagents into said assay plate structure to allow a fluid reagent mixture to be retained in each well, and   optical assessment means for measuring optical result of the reaction in each well.       
 
         [0056]    Preferably, the fluid handling means and the optical assessment means are automated. 
         [0057]    According to a sixth aspect of the present invention there is provided an assay plate structure for use in conducting optical assays of a fluid analyte, the plate structure comprising:
       a disc for rotation about a central axis, the disc having upper and lower plates and a plurality of substantially radially extending walls disposed between the plate, wherein said walls sub-divide the disc into a plurality of disc sectors; and   a plurality of disc inserts arranged to be received by respective disk sectors and to be retained therein,   the structure further having a plurality of openings through the upper surface, at least one opening above each disc sector for introducing a liquid analyte into the sector space between the plate and the disc insert.       
 
         [0061]    Preferably, the disc further comprises a lower plate, spaced apart from said upper plate by said radially extending walls. More preferably, the upper and lower plates are circular. 
         [0062]    Preferably, the upper surface of each disc insert and the opposed surface of the plate are substantially planar, and, more preferably, are in a closely spaced arrangement. 
         [0063]    Preferably, a vent opening is provided for each disc segment around the periphery thereof, between the radially outer edge of the upper plate and each disc insert. 
     
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS  
         [0064]    These and other aspects of the present invention will now be described with reference to the accompanying drawings, in which: 
           [0065]      FIG. 1  is a diagrammatic representation of a multi-well assay plate structure according to a first embodiment of the present invention; 
           [0066]      FIGS. 2   a  to  2   c  illustrate the steps involved in filling the wells of the container of  FIG. 1 ; 
           [0067]      FIG. 2   d  is an enlarged detail of part of the structure of  FIGS. 2   a  to  2   c;    
           [0068]      FIG. 3  shows a multi-well assay plate structure according to a second embodiment of the present invention; 
           [0069]      FIG. 4   a  shows a third embodiment of a disc-style structure for conducting multi-tests; 
           [0070]      FIG. 4   b  shows an enlarged cross-sectional detail of  FIG. 4   a  to allow snap-fitting of the plates in the disc sectors; 
           [0071]      FIG. 4   c  is a fourth embodiment of a disc-style structure for conducting multi-tests; 
           [0072]      FIG. 4   d  shows a modification of the outer disc with hinged sectors and which is applicable to the previous embodiments; 
           [0073]      FIG. 5  depicts chemical/biochemical assay apparatus for conducting an assay on reactions carried out using the multi-well assay plate structures shown in  FIG. 3  or  FIGS. 4   a, b, c  and  d , and 
           [0074]      FIGS. 6   a  and  6   b  depict the data and graphs respectively of antigen/antibody biochemical assays carried out using the apparatus of  FIG. 5  on the assay plate shown in  FIGS. 4   a, b, c  and  d.    
       
    
    
     DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS 
       [0075]    Reference is first made to FIG. I which shows a multi-well assay plate, generally indicated by reference numeral  10 , having a box-like construction with a rectangular cross-section. The assay plate  10  comprises an upper plate  12 , a lower plate  14 , and side and rear spacers  16 , 18 , 20  all of which are made of a transparent polycarbonate. The front of the box, indicated generally by the reference numeral  22 , is open to the surrounding space. 
         [0076]    The spacers  16 , 18 , 20  are dimensioned to produce a space  21  of uniform spacing d between the opposed inner surfaces  12   a ,  14   a  of the upper and lower plates  12 , 14 . Spacing d is chosen such that a selected liquid is able to flow through the space  21  between the upper and lower plates  12 , 14  in a controlled manner by capillary or capillary-like action. Generally, d is less than 0.5 mm. 
         [0077]    A small opening  23  extends through the upper plate  12  to communicate the inner space  21  with the exterior space surrounding the container. Opening  23  is located close to the rear wall  20  in order to prevent air-locks forming in the container during filling as will be described in more detail below. 
         [0078]    A regular array of wells or depressions  24  are formed in the upper surface  14   a  of the lower plate  14 . Typically, the polycarbonate assay plate with wells  24  is produced by suitably molding the lower plate  14  or by etching or pressing. The wells  24  are 2 mm in diameter and 1 mm deep and typically have a volume of 5 μl and any suitable number of wells may be provided. The wells are spaced 4 mm apart (center to center). 
         [0079]      FIGS. 2   a  to  2   c  illustrate the process by which the wells  24  of the assay plate  10  are filled with a liquid analyte  25 . The end  26  of a syringe  28  containing the liquid analyte  25  is pressed into the opening  23  provided in the upper plate  12  of the container  10  ( FIG. 2   a ) so as to form an air-tight seal between the periphery of the syringe and the inner surface of the opening  23 . The plunger  30  of the syringe  28  is then depressed to force the liquid  25  through the opening  23  into the space  21  within the plate  10 . As best seen in  FIG. 2   b , due to the capillary or capillary like flow of liquid through the space  21 , the entire space  21  is filled and wells  24  are covered before liquid  25  begins to flow through the front open face  22  of the container  10 . When it is observed that all of the space  21  is filled and the wells  24  are covered with liquid, and preferably prior to liquid flowing out through the front face  22 , the plunger  30  of the syringe  28  is withdrawn. This action empties the space  21  of liquid, but results in the wells  24  being filled with liquid  25  as shown in  FIG. 2   c .  FIG. 2   d  shows an enlarged cross-sectional view through part of the assay plate structure and showing how liquid is retained in wells  24  up to the meniscus. As with the filling process, liquid flows from the space  21  in a controlled manner. No puddles or drops of liquid remain in the space  22 , other than in the wells  24 . 
         [0080]    It will be appreciated that prior to introducing the liquid analyte  25  into the space  21 , for example during the manufacture of the assay plate  10 , the wells  24  of the plate  10  may be coated with an appropriate reactant. For example, if it is desired to conduct antigen-antibody reactions, the wells  24  are coated with an antigen. The remainder of the surface  14   a  is coated with a blocking agent to prevent antigen and antibodies from binding to surface  14   a . Once the wells  24  have been filled with the liquid analyte  25 , any antibodies present in the liquid analyte  25  will bind with the antigens contained in the wells  24 . There is no binding of the antibodies to surface  14   a . If it is necessary to conduct a further reaction in the wells  24 , e.g. to bind a colored or fluorescent label to the bound antibodies or exposed antigens, it is possible to repeat the steps of  FIGS. 2   a  to  2   c  in order to introduce the labelled components into the wells  24 . Prior to introducing the labelled components, if it is necessary to rinse the wells  24  and the inner surfaces  12   a ,  14   a  of the plate  10 , this is again easily achieved by repeating steps  2   a  to  2   c  with the syringe  28  containing, for example, distilled water. 
         [0081]    There is illustrated in  FIG. 3  a second embodiment of the present invention which depicts a multi-well assay plate in the form of a disk  32  designed for use with a rotating scanning device having a CD player type format. One such device is described for example in WO96/09548. The disk  32  shown in  FIG. 3  comprises a pair of upper and lower circular plates  34 , 36  sandwiched together to provide a cylindrical space  38  therebetween. This space  38  is divided into eight sectors  40  by radially extending spacers  42 . A plurality of wells  44  are provided in each sector  40  (one set of which is shown in broken outline) by forming the upper surface  36   a  of the lower circular plate  36  as described with reference to  FIG. 1 . The wells  44  are of the same size and are spaced as for  FIG. 1 . 
         [0082]    Each sector  40  provides a chamber or space  46  which can be filled independently via openings  48  provided through the top surface of each sector  40 . The peripheral edge  50  of each sector  40  is open to the surrounding space to provide a vent for the sector  40  to allow liquid to flow through the space or chamber  46  by displacing air therefrom. 
         [0083]    In order to enable the disk  32  to be compatible with scanning devices such as are described in WO 96/09548, the upper and/or lower plates  34 , 36  are made of transparent polycarbonate to enable a light beam to be scanned across the disk surface. The disk  32  is provided with a central hole  52  to enable the disk  32  to be mounted on a rotatable shaft. 
         [0084]    As is described in WO 96/09548, one of the surfaces of the upper or lower plates  34 ,  36  may be provided with digitally encoded address information which can be read by the scanned light beam. This information may be encoded by way of “pits” and “lans” pressed or molded into one of the plates. This address information can be used to provide accurate location information on the part of the disk which is begin scanned by the light beam. 
         [0085]    There is shown in  FIG. 4  a third embodiment of a disk assay plate  54  which comprises upper and lower circular transparent polycarbonate plates  56 , 58  which are spaced apart by a number of radially extending spacer walls  60  to create a plurality of disk sectors  62 . The inner surfaces  56   a ,  58   a  of the circular plates  56 ,  58  are both planar. 
         [0086]    Each disk sector  62  is arranged to receive a sector plate insert  64  which is a transparent polycarbonate plate with a detachable handle  66  on the outer side to facilitate entry and removal of the plate insert  64  in the sector  62 . The plate insert  64  and spacer wall  60  have respective recesses/projections (not shown in the interest of clarity) which allow the assay plate  64  to be inserted only in the correct orientation. The plate  64  has a groove  68 , as shown in  FIG. 4   b  for example, which allows the inset to be snap-fitted over a projection  70  upstanding from plate  58  into the sector. The thickness of the sector insert plate  64  is marginally less than the spacing provided between the upper and lower plates  56 , 58  so that the plate insert  64  can be pressed/fitted into one of the disk sector  62  to define a liquid receiving chamber or space  73  between the upper surface  64   a  of the insert plate  64  and the lower surface  56   a  of the upper disk plate  56 . Openings  72  are provided through the upper plate  56  into each disk sector  64  whilst the space  70  between the radially outermost peripheral edge  74  of the insert plate  64  and the upper plate  56  provides a further vent or filling opening into the disk sector  62 . 
         [0087]    The surface  64   a  of the insert plate  64  is provided with a plurality of wells  76  as described with respect to  FIG. 1 . The wells are 2 mm in diameter, 1 mm in depth and 4 mm apart (spaced between centers). These wells are filled by introducing liquid into the disk sector  64  through the upper opening  72  to fill space  70  and subsequently withdrawing the liquid through the same opening as previously described. 
         [0088]    Reference is now made to  FIG. 5  of the drawings which depicts assay apparatus for conducting an assay on reactions carried out using the assay plate structures of the already described embodiments. However, for convenience, the assay apparatus will be described in combination with the preferred embodiment shown in  FIGS. 4   a, b  with like numerals referring to like parts. 
         [0089]    In this case the plate  54  is mounted on a shaft  74  carried by a turntable  77 . The apparatus includes a suitable automatic fluid filling/withdrawal system, generally indicated by reference numeral  80 , which operates a syringe  82  to dispense/retrieve fluid from a reservoir  84  via the openings  72  into the space  70  between the plate surface  56   a  and the surface  64   a  of each sector plate  64 . The fluid can of course be dispensed and retrieved manually if desired. This is achieved for each sector by rotating the disk plate  54  to a suitable position to allow fluid filling/withdrawal. It will be appreciated that the plates are pre-prepared with various reagents, e.g. antigens, and they are inserted in the appropriate wells  76 , as described with reference to  FIGS. 4   a ,  4   b . The plates are first flooded with fluid carrying antibodies and withdrawal of the fluid leaves the antibody/antigen reagents filling the wells  76  resulting in a reaction. 
         [0090]    The following example of an assay within the embodiment shown in  FIG. 4   b  is described to provide a better understanding of the steps involved: 
       Multi-Antigen Elisa Using Sectors 
       [0091]    1. The underside of upper surface ( 56   a ) of is coated with silicone spray to aid fluid movement. Sector plates  64  are also coated including wells  76 . Any excess silicone is removed. 
         [0092]    2. Sectors wells  76  are loaded by hand with a panel of seven antigens—Human Serum Albumin, Antitrypsin, Macroglobulin, Antithrombin III, Catalase, Antichymotrypsin and Plasminogen at a concentration of 20 μg/ml in PBS and a volume of 2 μl/well. Control wells contain PBS only. Antigens can be arranged in blocks of the same on the sector plate  64  in a series giving a panel of tests evenly distributed over the sector. Incubate at room temperature for 15 minutes. 
         [0093]    3. Wash with 0.05% PBS-Tween using flood/fill technique—1 ml is flooded across the sector plate via holes  72  in the top plate using a 1 ml pipette. This pipetted up and down three times then withdrawn and the washing discarded. This repeated a further three times to ensure complete washing. 
         [0094]    4. Blocking is carried out to prevent reactions occurring other than at well sites with 50 mg/ml Bovine Serum Albumin (BSA) (in PBS) using flood/fill. 1 ml of BSA/PBS is flooded across the sector, pipetted up and down three times, withdrawn and discarded. This allows all wells  76  to be filled simultaneously. Incubate for 15 minutes at room temperature. 
         [0095]    5. Wash as before. 
         [0096]    6. Primary antibodies are applied to the sector plate  64  as a mixture using flood/fill with each individual antibody at the following concentrations: anti-Human Serum Albumin 1/1000, anti-Antitrypsin 1/2000, anti-Macroglobulin 1/2000, anti-Antithrombin III 1/1000, anti-Catalase 1,1000, anti-Antichymotrypsin 1/1000, anti-Plasminogen 1/1000. Antibodies are diluted in 0.5 mg/ml BSA/PBS. Incubate for 10 minutes at room temperature. 
         [0097]    7. Wash as before. 
         [0098]    8. Second antibody is Amdex anti-IgG (peroxidase conjugate) at a concentration of 1/1000 in 0.5 mg/ml BSA/PBS. After washing this is applied to the sector using flood/fill. Incubate at room temperature for 10 minutes. 
         [0099]    9. Wash as before. 
         [0100]    10. The substrate is insoluble Tetramethylbenzidine (TMB). This reacts with the peroxidase on the second antibody to produce an intense blue color. After washing this is applied to the sector plate  64  by flood/fill but is left flooded across the sector plate  64  after pipetting up and down several times. Incubate for 10 minutes at room temperature. 
         [0101]    11. Remove TMB and discard. Wash out the wells with distilled water four times by flood/fill. A blue precipitate will be evident in wells with a positive reaction. No color is produced in negative wells. Store sections in dark as TMB will slowly fade in daylight. 
         [0102]    The data for the above assay is shown in  FIG. 6   a  and is graphically represented in  FIG. 6   b  which is reproducible and is representative of a large number of experiments ( 712 ). 
         [0103]    It will be seen that there is a significant measurable change for each antibody/antigen reaction compared with the background level. The reaction results in an optical change, from transparent to colored (blue) and which is measured using an optical detector which measures light transmissivity through the disk and wells. In this case optical assessment was carried out using the apparatus as shown in  FIG. 5  by locating the plate  64  in a light transmissive microscope  80  (Zeiss Axiophot fitted with a JVC video camera  83  (Model No. TK-1280E)) and sensing the change in optical signal. The output of the video camera is connected to Macintosh IICx  85  with video frame capture. The results can be displayed via the Mac display  87  or a hard copy provided by printer  86 . Analysis was carried out by measuring mean grayscale values in center of wells quantified by NIH Image software. Background levels taken from sectors which had not been exposed to immunochemicals or chromogen were subtracted from all experimental wells. Experimental wells contained array or seven separate antigens listed above. In addition, experimental controls were carried out in which specific antigen was omitted wells and wells exposed to the same regime of blocking, antibody binding and exposure to chromogenic substrate. The average reading from these experimental controls minus mean reading from the sector alone was defined as the background level of staining. Experimental readings from the seven specific antigens providing signals of approximately five to six times greater than this background. It will be observed that there is no cross-contamination between wells  76  because of the efficiency of withdrawal and because the substrate in this case is insoluble. However, this assay would also work satisfactorily for soluble substrates because of fluid withdrawal from the sector plate  64  leaving fluid in the wells  76  only, not on surface  64   a.    
         [0104]    In a modification, if it was unnecessary to withdraw all of the liquid to leave a film on surface  64 , the assay would still work with an insoluble substrate in each well; cross-contamination would still not occur. However, this arrangement would be unsatisfactory for soluble substrates in the wells as the film could cause dispersal to other locations and provide contamination of other wells. 
         [0105]    With the embodiment shown in  FIGS. 4   a ,  4   b  the disk sector plate  54  is more suitable for conducting a variety of different assays, e.g. antigen/antibody assays for different patients, i.e. one patient/sector. 
         [0106]    It will be appreciated that modifications may be made to the above described embodiments without departing from the scope of the present invention. For example, the opening through which the liquid analyte is introduced may be provided through the lower plate of the multi-well container. More than one opening, can be used for faster flooding. This opening may be arranged to receive the tip of a syringe needle. The vent opening may also be provided in any one of the walls of the container although it is preferably provided in a peripheral wall. The opening  22  may be provided by a single opening  22  or by a series of openings or vents as shown in  FIG. 4   d  for example. A laser may be used with CD optics instead of the microscope and video camera for the embodiment of  FIG. 4 . The top plate in the embodiment of  FIGS. 3 and 4  may be snap-fitted to the lower plate and may be snap-fitted onto a CD base plate which would receive sections and provide the advantage of positioned information. As shown in  FIG. 4   c  the upper planar surface  56  can have sector covers connected to a lower surface or central boss by a hinge, for example integrated living hinge  90  at the inner radius to allow each disk sector  62  to be pivotably raised and lowered and allow sector plates  64  to be inserted into each sector. The well size and spacing may be varied as required, for example the wells could be 3 mm in diameter; 1.5 mm apart and spaced 5.5 mm between center. The exact size and spacing is a matter of choice consistent with the requirement that fluid is retained in the wells after withdrawal as described above. However, the wells could also be filled during flooding of the space depending on the well size, type of plastic and fluid properties. However, liquid will still be retained in the wells upon withdrawal of the liquid. Also, the structure and inserts made may be of any suitable optically transmissive plastic, such as polystyrene or perspex™. The handle  66  may be integrated with or detachable from plate  64 . As shown in  FIG. 4   a  the radially extending ribs may have radial shoulders  92  to define a recess  94  for receiving the plate  64  also defining the spacing height between the surface  64   a  of the plate  64  and the underside  56   a  for receiving the liquid. Suitable materials may be used to coat the interior of the sectors to aid fluid movement as described with reference to silicone above. This may be applied to the underside of the top surface and to the top surface of the plate as for the other embodiments. Suitable materials may be used to increase the hydrophobicity of liquid across the sector and hydrophilicity to and movement of liquid into the desired location, e.g. wells. The wells may be coated with a suitable optical reflective material to enhance the reflection of light and observation of reactions occurring within the wells and, similarly, lenses  90  may be located in the top or bottom light transmissive plates  12  and  14  as seen in  FIG. 8 , to improve optical assessment of the reaction. These lenses may be moulded into the upper or lower plates of the exemplary embodiments during the manufacture as is well known in plastic molding processes. Separate optical elements may be used instead, if appropriate. 
         [0107]    In a modification to the embodiments described, the wells are absent from the upper surface of the plate and that plate retains its planar surface to enable a thin, uniform layer of liquid to be introduced into the space between the upper disk plate and the insert plate. An insoluble substrate with reagent or reagents (e.g. an antigen) may be applied directly to the planar surface of the insert plate by for example applying spots of reagent thereto. 
         [0108]    For certain applications, it may be appropriate to provide each insert with a lid which can be slid into the space between the insert and the upper plate  22  of the disk following filling of the wells. The lower surface of the lid may be arranged to be flush with the surface of the insert so as to close off each well. This prevents liquid from being thrown out of the wells during spinning of the disk during automated reading and analysis. The invention has use in immunoassay applications including tests for sexually transmitted diseases, parasites, allergens, cancer markers and cardiac markers, either in laboratories or at point-of-care locations, for example medical practitioners offices or the like. Other applications of the invention are in chemical and biochemical assays. Examples of such assays include immunoassay, clinical biochemistry tests, nucleic acid analysis and receptor ligand interactions. Examples of clinical biochemistry uses would be in measurement of serum analytes such as glucose, urea, creatinine and enzymes such as alkaline phosphatase. Immunoassay application include tests designed to detect infectious organisms, viruses, parasites as well as endogenous analytes such as circulating hormone levels and cancer markers. Examples of chemical analysis include measure of phosphate and nitrate levels in water, environmental and industrial monitoring including potable and waste water and process monitoring. The system could be used in a variety of settings including clinical laboratories, doctor&#39;s and veterinary surgeries as well as industrial and research laboratories.