Abstract:
A device for assisting in fluid mixing within microfluidic sized structures. Chemicals and other biological specimens are exposed to a small volume of reagent, and said reagent is delivered to said specimens by a novel mixing technique, thus shortening overall process time.

Description:
CROSS-REFERENCE TO RELATED APPLICATIONS  
       [0001]    This patent application claims benefit from U.S. Provisional Application Ser. No. 60/213,865, filed Jun. 23, 2000, which application is incorporated herein by reference. 
     
    
     
       BACKGROUND OF THE INVENTION  
         [0002]    1. Field of the Invention  
           [0003]    This invention relates generally to mixing of fluids and, in particular, to the mixing of small volumes of fluids that are dispersed over a relatively large area. Such mixing is required, for example, when a small quantity of reagent is to be distributed uniformly over a microscope slide. This is desired when spotted microarrays are to be exposed to various reagents.  
           [0004]    2. Description of the Prior Art  
           [0005]    Spotted microarrays are currently in use for various applications. In most cases, a spotted microarray comprises a glass slide of roughly 1″ by 3″, onto which several hundred to several thousand spots are deposited. These spots typically contain genetic material or other material of biological interest.  
           [0006]    Microarrays are currently exposed to reagents by dipping them into a fairly large volume of fluids. Given the sometimes very high costs of the reagents, it would be desirable to reduce the volume required for exposing the microarrays.  
           [0007]    Microfluidic devices have recently become popular for performing analytical testing. Using tools developed by the semiconductor industry to miniaturize electronics, it has become possible to fabricate intricate fluid systems which can be inexpensively mass produced. Systems have been developed to perform a variety of analytical techniques for the acquisition of information for the medical field.  
           [0008]    Microfluidic devices may be constructed in a multi-layer laminated structure where each layer has channels and structures fabricated from a laminate material to form microscale voids or channels where fluids flow. A microscale channel is generally defined as a fluid passage which has at least one internal cross-sectional dimension that is less than 500 μm and typically between about 0.1 μm and about 500 μm. The control and pumping of fluids through these channels is affected by either external pressurized fluid forced into the laminate, or by structures located within the laminate.  
           [0009]    A microfluidic device can be constructed that mates with a spotted microarray such that the microarray forms the bottom of a channel that is as wide and as long as the microarray, but has a depth of a microfluidic dimension. Microfluidic devices are defined as having at least one dimension in the range of 1-1000 micrometers. Typically, such a device would have a channel depth of 100 micrometers.  
           [0010]    Providing such a device does solve the problem of reducing the reagent volume requirement, but creates another problem: all fluid flow in such a channel is laminar, which implies that, when fluid flows into a such a channel, no mixing other than by diffusion of particles occurs. Particle diffusion is a slow process, depending on the particle size and other fluid parameters, and it can take several hours for fluid particles to diffuse a distance of a few millimeters. Therefore, reactions between the chemicals immobilized in a spot, and those contained in the reagent solution, are rate-limited by the diffusion of reagent particles to the spot. This significantly slows down the reaction, and therefore the required process time for spotted arrays. This invention provides a device and method for moving fluid in such a channel such that each spot is periodically or continuously exposed to a fresh, unreacted portion of the reagent fluid such that the chemical reaction is no longer diffusion-limited, and the overall process time is reduced.  
         SUMMARY OF THE INVENTION  
         [0011]    It is therefore an object of the present invention to provide a method and a device for mixing fluids in wide channels that have a depth of a microfluidic dimension.  
           [0012]    It is a further object of the present invention to provide a method and a device for exposing chemicals that are immobilized on a slide to a small volume of reagent while preventing the reaction from becoming diffusion-limited.  
           [0013]    These and other objects of the present invention will be more readily apparent in the description and drawings that follow. 
       
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0014]    [0014]FIG. 1 is a top view of a mixing process used in the prior art;  
         [0015]    [0015]FIG. 2 is a top view of a slide prepared according to the present invention;  
         [0016]    [0016]FIG. 3 is a top view of a cover slide for use with the slide shown in FIG. 2;  
         [0017]    [0017]FIG. 4 is a top view of the glass slides of FIGS. 2 and 3 during the mixing process;  
         [0018]    [0018]FIG. 5 is a top view of an alternative embodiment of the present invention;  
         [0019]    [0019]FIG. 6 is a front view of an alternative embodiment of the present invention which uses syringe pumps to assist in fluid mixing;  
         [0020]    [0020]FIG. 7. is a front view of an alternative embodiment of the present invention which uses a bubble pump to assist in fluid mixing;  
         [0021]    [0021]FIG. 8 is a front view of the device shown in FIG. 7 in rotation;  
         [0022]    [0022]FIG. 9 is a front view of another alternative embodiment of the present invention which uses rotation of the entire device to assist in mixing; and  
         [0023]    [0023]FIG. 10 is a front view of the device shown in FIG. 9 showing different locations during rotation of the device. 
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS  
       [0024]    [0024]FIG. 1 is a representation of the current procedure which is commonly used in laboratories. Referring now to FIG. 1, there is shown a microscope slide  10  containing an array  12  of sample microdots. A reacting liquid is placed on slide  10  covering array  12 , and then a cover slip  14  is placed on slide  10 , covering array  12 . Slide  10  is processed with a heat cycler, and slide  10  is then set aside so that diffusion can take place, as no active mixing occurs during this procedure. Diffusion of the reacting liquid can take as long as 24 hours, and often longer, as the reaction is diffusion-limited. The incubation period for this process can often be very long.  
         [0025]    A novel method for performing microfluidic fluid mixing is shown in FIGS.  2 - 4 . Referring now to FIG. 2, a circular slide  20  is shown containing an array  22  of microdots, while the center area  24  of slide  20  contains no microdots, as array  22  comprises a toroidal shape on slide  10 . A toroidal cover slide  26 , shown in FIG. 3 is also circular in shape, and has a circular aperture  28  located in the central portion of cover slide  26  which aperture corresponds to area  24  of slide  20 .  
         [0026]    To begin the reaction process, a reacting liquid is placed in aperture  28  of cover slide  26 . The liquid will wick under cover slide  26  by capillary action. Cover slide is then rotated in the direction shown by arrow A in FIG. 4. This motion causes the liquid to be completely across the array  22  of microdots, allowing the reaction between the microdots and the reacting liquid. Surface tension at the edges of slide  20  and cover slide  26  will contain the reacting fluid between the slides. The result of this process is a shortened incubation period.  
         [0027]    [0027]FIG. 5 shows an alternative embodiment of the invention taught in FIGS.  2 - 4  using different geometries. A rectangular microscope slide  30  is shown having an area  32  in which an array  34  of microdots are located, leaving an area  36  in which no microdots are found. A circular glass slide  38  is initially positioned in area  36 .  
         [0028]    The mixing process begins as a reacting fluid is added to array  34  and circular slide  38  spins in the direction shown by arrow B while slide  38  moves across array  34  and oscillates back and forth across slide  30 . Rotating slide  38  causes local Couette flow as it passes across the microdots in array  34  on slide  30 . An external container  40  is used to container slide  30  to inhibit evaporation.  
         [0029]    Another embodiment of the present invention is shown in FIG. 6 using a pair of syringe pumps. A glass microscope slide  50  having an array  52  of microdots positioned thereon has a pair of syringe pumps  54 ,  56  positioned at each end. A cover slide  58  is located above array  52  over the microdots. Reacting fluid is loaded into syringes  54 ,  56  and each syringe is operated 180° out of phase such that fluid is expelled from one syringe as it is taken up by the other syringe. This motion causes a Poiseuille flow across array  52  of microdots.  
         [0030]    Another embodiment of the present invention is shown in FIGS. 6 and 7, which embodiment operates as a bubble pump. A glass microscope slide  60  having an array  62  of microdots applied to the upper surface is covered with a reacting fluid  64  within an enclosure  66 . A slide  68  is located within enclosure  66  covering array  62 . Fluid  64  fills enclosure  66  such that an air bubble  70  is trapped within enclosure  66  above cover slide  68 .  
         [0031]    Another version of this embodiment uses a second fluid which is substantially immiscible and has a different density than said reacting fluid. The second fluid may contain magnetic particles or may have magnetic properties. The second fluid is then oscillated across array  62  by use of a magnetic field, such that the reacting fluid is also moved across array  66 . The same result may be accomplished by inserting magnetic particles into the reacting fluid.  
         [0032]    Enclosure  66  is then oscillated about a pivot point  72  with a rocking motion indicated by arrow C. The range of rotation is preferably limited to approximately 45° in the counterclockwise direction to 45° in the clockwise direction. As assembly  59  is rotated about point  72 , bubble  70  trapped within fluid  64  in enclosure  66  moves from end to end moves to the highest point, as can be clearly seen in FIG. 7, due to the air density being less than the fluid density. This gravity-induced motion will move fluid  64  below cover slide  68  back and forth across array  62 .  
         [0033]    An additional embodiment showing the present invention is shown in FIGS. 9 and 10. In this embodiment, a glass microscope slide  80  contains an array  82  of microdots positioned on the upper surface. Array  82  is covered with a processing liquid and is then covered by a cover slide  84 .  
         [0034]    Note that slide  84  only covers the area of slide  80  where array  82  is located. Slide  80  is then moved in a circular pattern without any movement of cover slide  84 . Several positions of slide  80  are shown in FIG. 10 as  84   a ,  84   b ,  84   c , and  84   d . This circular translation of slide  84  without rotation of cover slide  84  creates a form of Couette flow in the liquid covering array  82  between slide  84  and  82 . This flow mixes the fluid and brings chemical constituents contained in the liquid closer to the microdots in array  82  so that the diffusion path between the constituents within the liquid and the microdots is reduced, thus speeding up the reaction rate and reducing assembly time.  
         [0035]    While the present invention has been shown and described in terms of several preferred embodiments thereof, it will be understood that this invention is not limited to these particular embodiments and that many changes and modifications may be made without departing from the true spirit and scope of the invention as defined in the appended claims.