Patent Publication Number: US-2003235519-A1

Title: Protein crystallography hanging drop lid that individually covers each of the wells in a microplate

Description:
BACKGROUND OF THE INVENTION  
       [0001] 1. Field of the Invention  
       [0002] The present invention relates in general to the biotechnology field and, in particular, to a protein crystallography hanging drop lid designed to individually cover each of the wells in a microplate and methods for fabricating and using the protein crystallography hanging drop lid.  
       [0003] 2. Description of Related Art  
       [0004] Today biochemical studies associated with growing protein crystals and other biological crystals are carried out on a large scale in both industry and academia. As such, it is desirable to have an apparatus that allows researchers to perform these studies in a convenient and inexpensive fashion. Because they are relatively easy to handle and low in cost, microplates are often used in these studies. And, if the study involves growing protein crystals via a hanging drop vapor diffusion process, then the wells of a microplate are often covered with slides or a lid having one or more drops of a protein solution and a reagent solution hanging therefrom which turn into the protein crystals. In particular, the drops hanging from the bottom side of the slides or lid turn into protein crystals by interacting via a vapor diffusion process with a higher concentrated reagent solution located within each well of the microplate. However, the traditional slides or lid used to grow protein crystals in this manner have several drawbacks which are described in greater detail below with reference to FIGS.  1 - 3 .  
       [0005] Referring to FIGS.  1 A- 1 B (PRIOR ART), there are illustrated different views of one set of traditional slides  100  designed to cover the wells  104  in a microplate  102 . Each slide  100  typically has a circular shape and is sized to fit over one of the wells  104  in the microplate  102 . And, each well  104  includes a rim  106 , sidewalls  108  and a bottom  110  (see FIG. 1B). The wells  104  are generally arranged in a matrix of mutually perpendicular rows and columns. For example, the microplate  102  can include a matrix of wells  104  having dimensions of 4×6 (24 wells), 8×12 (96 wells) and 16×24 (384 wells). The microplate  102  shown includes an array of ninety-six wells  104 .  
       [0006] To grow a protein crystal on the bottom side of one slide  100 , the researcher applies a bead of grease  112  (e.g., high vacuum grease) along the rim  106  of one of the wells  104 . Typically, the researcher would leave a small opening such as  2 mm between the start and end of the bead of grease  112 . The researcher then pipets a small amount (e.g., 1.0 millimeter) of a reagent solution  114  into the well  104 . One or more drops  116  (only one shown) including a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter) that can be taken from the well  104  are then pipetted onto a bottom side of the slide  100 . Thereafter, the researcher inverts the slide  100  so that the drop  116  is hanging down from the slide  100  and then positions and places the slide  100  onto the grease  112  around the well  104 . To relieve the air pressure within the well  104 , the researcher presses the slide  100  down onto the grease  112  and twists the slide  100  to close the small opening in the grease  112 . This process is then completed for each well  104  in the microplate  102 . Unfortunately, there are a number of disadvantages associated with using the slides  100  and the microplate  102 . First, the researcher must work with messy grease  112  and possibly spend a lot of time applying the grease  112  to the rims  106  of each well  104 . Secondly, the researcher must work with and handle a large number of relatively small slides  100  to utilize all of the wells  104  in the microplate  102 . Thirdly, the slides  100  and the grease  112  are expensive.  
       [0007] Referring to FIGS.  2 A- 2 B (PRIOR ART), there are illustrated different views of another set of traditional slides  200  designed to cover the wells  204  in a microplate  202 . Each slide  200  typically has a circular shape and is sized to be placed on a ledge  203  in one of the wells  204  in the microplate  202 . And, each well  204  includes a rim  206 , sidewalls  208  and a bottom  210 . The wells  204  are generally arranged in a matrix of mutually perpendicular rows and columns. For example, the microplate  202  can include a matrix of wells  204  having dimensions of 4×6 (24 wells), 8×12 (96 wells) and 16×24 (384 wells). The microplate  202  shown includes an array of ninety-six wells  204 .  
       [0008] To grow a protein crystal on the bottom side of one slide  200 , the researcher pipets a small amount (e.g., 1.0 millimeter) of a reagent solution  214  into the well  204 . One or more drops  216  (only one shown) including a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter) that can be taken from the well  204  are then pipetted onto a bottom side of the slide  200 . Thereafter, the researcher inverts the slide  200  so that the drop  216  is hanging down from the slide  200  and then positions and places the slide  200  onto the ledge  203  within the well  204 . After, this process is completed for each well  204  in the microplate  202 , then the researcher places one or more strips of tape  218  (only shown in FIG. 2B) over the top of microplate  202 . Unfortunately, there are a number of disadvantages associated with using the slides  200  and the microplate  202 . First, the researcher must work with and handle a large number of relatively small slides  200  to utilize all of the wells  204  in the microplate  202 . Secondly, the researcher must cut the tape  218  in order to have access to anyone of the slides  200  located within a particular well  204 . Thirdly, the slides  200  are expensive.  
       [0009] Referring to FIGS.  3 A- 3 B (PRIOR ART), there are illustrated different views of a traditional lid  300  designed to cover the wells  312  in a microplate  302 . The lid  300  includes a rigid frame  304  that supports a filter membrane  306  on which there is placed a hydrophobic mask  308  all of which are protected by a removable cover  310  (see exploded view in FIG. 3B). The lid  300  is sized to fit over all of the wells  312  in the microplate  302 . And, each well  312  includes a rim  314 , sidewalls  316  and a bottom  318 . The wells  312  are generally arranged in a matrix of mutually perpendicular rows and columns. For example, the microplate  302  can include a matrix of wells  312  having dimensions of 4×6 (24 wells), 8×12 (96 wells) and 16×24 (384 wells). The microplate  302  shown includes an array of ninety-six wells  312 .  
       [0010] To grow a group of protein crystals on top of the hydrophobic mask  308  of the lid  300 , the researcher applies a bead of grease  320  (e.g., high vacuum grease) on the rims  314  of the wells  312  in the event the wells  312  are not pre-greased. The researcher then pipets a small amount (e.g., 1.0 millimeter) of a reagent solution  322  into each well  312 . One or more drops  324  (eight drops  324  are shown) including a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter) that can be taken from the well  104  are then pipetted onto the hydrophobic mask  308  of the lid  300 . Thereafter, the researcher positions the lid  300  over the microplate  302  and then pushes the lid  300  down onto the grease  322  located around each well  312 . The lid  300  can have holes  326  formed in the frame  304 . And, the microplate  302  can have pins  328  extending up therefrom which fit into the holes  326  in the frame  304  to assure that the lid  300  is properly aligned with the microplate  302 . Unfortunately, there are a number of disadvantages associated with using the lid  300  and the microplate  302 . First, the researcher must work with messy grease  320  and possibly spend a lot of time applying the grease  320  to the rims  314  of the wells  312 . Secondly, the filter membrane  306  and hydrophobic mask  308  of the lid  300  are very fragile and can easily break. Thirdly, the lid  300  is very expensive.  
       [0011] Accordingly, there is and has been a need for a cost effective and user-friendly lid that can be used with a microplate to help a researcher perform protein crystallization studies. This need and other needs are satisfied by the lid and the methods of the present invention.  
       BRIEF DESCRIPTION OF THE INVENTION  
       [0012] The present invention includes a lid that individually covers each of the wells in a microplate and methods for fabricating and using the lid. The lid includes a series of downwardly protruding necks each of which is sized to fit and seal against one of the wells formed within the microplate. And, each neck has a surface covered with a rubber-like substance that ensures a relatively tight seal with each well. In operation, the lid and microplate are preferably used together to grow protein crystals using a hanging drop vapor diffusion crystallization process. 
     
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
     [0013] A more complete understanding of the present invention may be had by reference to the following detailed description when taken in conjunction with the accompanying drawings wherein:  
     [0014] FIGS.  1 A- 1 B (PRIOR ART) illustrates different views of one set of traditional slides made by Hampton Research Corporation that are designed to cover the wells in a microplate;  
     [0015]FIG. 2A- 2 B (PRIOR ART) illustrates different views of another set of traditional slides made by Hampton Research Corporation that are designed to cover the wells in a microplate;  
     [0016] FIGS.  3 A- 3 B (PRIOR ART) illustrates different views of a traditional lid made by Neuro Probe Incorporated that is designed to cover the wells in a microplate;  
     [0017] FIGS.  4 A- 4 H are different views of a first embodiment of a lid designed to cover the wells of a microplate in accordance with the present invention;  
     [0018] FIGS.  5 A- 5 H are different views illustrating a second embodiment of a lid designed to cover the wells of a microplate in accordance with the present invention;  
     [0019] FIGS.  6 A- 6 H are different views illustrating a third embodiment of a lid designed to cover the wells of a microplate in accordance with the present invention;  
     [0020] FIGS.  7 A- 7 C are partial cross-sectional side views of different microfluidic channels that can be incorporated within anyone of the lids shown in FIGS.  4 - 6 ;  
     [0021]FIG. 8 is a flowchart illustrating the steps of a preferred method for using a lid and a microplate in accordance with the present invention; and  
     [0022]FIG. 9 is a flowchart illustrating the steps of a preferred method for fabricating a lid in accordance with the present invention. 
    
    
     DETAILED DESCRIPTION OF THE DRAWINGS  
     [0023] Referring to FIGS.  4 - 9 , there are disclosed in accordance with the present invention several embodiments of a lid  400 ,  500  and  600  designed to cover a microplate  402 ,  502  and  602  and methods  800  and  900  for fabricating and using the lid. Although the lid  400 ,  500  and  600  and the microplate  402 ,  502  and  602  are described as being used to grow protein crystals using a hanging drop vapor diffusion crystallization process, it should be understood that the lid and microplate are not limited to this application. Instead, the lid  400 ,  500  and  600  can be used with the microplate  402 ,  502  and  602  to perform a wide variety of applications including one where the lid simply ensures that a solution remains within the wells of the microplate. Accordingly, the lid  400 ,  500  and  600  and the methods  800  and  900  for fabricating and using the lid should not be construed in a limited manner.  
     [0024] Referring to FIGS.  4 A- 4 H, there are illustrated different views of a first embodiment of a lid  400  designed to cover the individual wells  404  of a microplate  402 . The microplate  402  includes an array of wells  404  each of which has a rim  406 , sidewalls  408  and a bottom  410 . The wells  404  are generally arranged in a matrix of mutually perpendicular rows and columns. For example, the microplate  402  can include a matrix of wells  404  having dimensions of 4×6 (24 wells), 8×12 (96 wells) and 16×24 (384 wells). The microplate  402  shown includes an array of ninety-six wells  404 .  
     [0025] The lid  400  (e.g., protein crystallography hanging drop lid  400 ) is sized and configured to individually cover each of the wells  404  in the microplate  402 . In particular, the lid  400  includes a frame  412  having formed therein a series of downwardly protruding necks  414  (see cross-sectional side view in FIG. 4B and partial bottom view in FIG. 4C). Each neck  414  is designed to fit and seal against one of the wells  404  in the microplate  402 . In one embodiment shown in FIG. 4D, each neck  414  is designed to fit and seal against an inside edge of the rim  406  extending from one of the wells  404  in the microplate  402  (see also FIG. 4B). In another embodiment shown in FIG. 4E, each neck  414  is designed to fit and seal against an outside edge of the rim  406  extending from one of the wells  404  in the microplate  402 .  
     [0026] To ensure a relatively tight seal between each neck  414  and each well  404 , all or a portion of the bottom side of the frame  412  and the outer surface of each neck  414  are covered with a rubber-like substance  422  (shown as shaded area). In the preferred embodiment, the rubber-like substance  422  is a thermoplastic elastomer. However, any rubber-like substance  422  can be used that has gasket-like characteristics with the appropriate flexure and friction properties that can ensure a consistently tight seal around the wells  404 . The relatively tight seal between each neck  414  and each well  404 , should be maintained regardless of the different dimensional tolerances of the wells  404 , the hand assembly of the lid  400  to the microplate  402  and the movement of the lid  400  and microplate  402 .  
     [0027] Each neck  414  has a transparent window  424  which is part of the frame  412  that enables a user to see inside each well  404  when the lid  400  is attached to the microplate  402 . The window  424  is transparent because the frame  412  is preferably made from a clear substance such as polystyrene, cyclic olefin or polypropylene and the rubber-like substance  422  is a colored substance such as a colored thermoplastic elastomer. Alternatively, the rubber-like substance  422  could be clear and does not need to be colored. Each window  424  has a bottom side  426  that is located within an inner surface of the neck  414 . In the embodiments shown in FIGS.  4 D- 4 E, each window  424  has a flat bottom side  426  that is flush with a top of the rim  406  of the well  404 . In another embodiment shown in FIG. 4F, each window  424  has a flat bottom side  426  that is located within the well  404 . In yet another embodiment shown in FIG. 4G, each window  424  has a flat bottom side  426  that is located above the well  404 . In still yet another embodiment shown in FIG. 4H, each window  424  has a concaved bottom side  426 . Of course, each window  424  may have a bottom side  426  with other shapes and locations other than those described above with respect to FIGS.  4 D- 4 H.  
     [0028] To grow one or more protein crystals using the lid  400  and microplate  402 , the researcher pipets a small amount (e.g., 1.0 millimeter) of a reagent solution  430  into each of the wells  404  (see, FIGS.  4 D- 4 H). One or more drops  432  (only one shown) including a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter) that can be taken from the wells  404  are then pipetted onto the bottom sides  426  of the necks  414 . Thereafter, the researcher inverts the lid  400  so the drops  432  hang down from the lid  400  and then positions, pushes-down and secures the lid  400  onto the microplate  402 . In this position, the drops  432  via a vapor diffusion process turn into protein crystals by interacting with the higher concentrated reagent solution  430  located within the wells  404  of the microplate  402 . Instead of having the researcher handle the lid  400 , the lid  400  can have a footprint which makes it capable of being handled by a robotic handling system (not shown).  
     [0029] Referring to FIGS.  5 A- 5 H, there are illustrated different views of a second embodiment of a lid  500  designed to cover the individual wells  504  of a microplate  502 . The microplate  502  includes an array of wells  504  each of which has a rim  506 , sidewalls  508  and a bottom  510 . The wells  504  are generally arranged in a matrix of mutually perpendicular rows and columns. For example, the microplate  502  can include a matrix of wells  504  having dimensions of 4×6 (24 wells), 8×12 (96 wells) and 16×24 (384 wells). The microplate  502  shown includes an array of ninety-six wells  504 .  
     [0030] The lid  500  (e.g., protein crystallography hanging drop lid  500 ) is sized and configured to individually cover each of the wells  504  in the microplate  502 . In particular, the lid  500  includes a frame  512  having formed therein a series of downwardly protruding necks  514  (see cross-sectional side view in FIG. 5B and partial bottom view in FIG. 5C). Each neck  514  is designed to fit and seal against one of the wells  504  in the microplate  502 . In one embodiment shown in FIG. 5D, each neck  514  is designed to fit and seal against an inside edge of the rim  506  extending from one of the wells  504  in the microplate  502  (see also FIG. 5B). In another embodiment shown in FIG. 5E, each neck  514  is designed to fit and seal against an outside edge of the rim  506  extending from one of the wells  504  in the microplate  502 .  
     [0031] To ensure a relatively tight seal between each neck  514  and each well  504 , all or a portion of the bottom side and a portion of the top side of the frame  512  and the outer surface of each neck  514  are covered with a rubber-like substance  522  (shown as shaded area). In the second embodiment, the rubber-like substance  522  is also located on the top side of the frame  512  except where the windows  524  are located (compare to lid  400  in FIG. 4B). In the preferred embodiment, the rubber-like substance  522  is a thermoplastic elastomer. However, any rubber-like substance  522  can be used that has gasket-like characteristics with the appropriate flexure and friction properties that can ensure a consistently tight seal around the wells  504 . The relatively tight seal between each neck  514  and each well  504 , should be maintained regardless of the different dimensional tolerances of the wells  504 , the hand assembly of the lid  500  to the microplate  502  and the movement of the lid  500  and microplate  502 .  
     [0032] Each neck  514  has a transparent window  524  which is part of the frame  512  that enables a user to see inside each well  504  when the lid  500  is attached to the microplate  502 . The window  524  is transparent because the frame  512  is preferably made from a clear substance such as polystyrene, cyclic olefin or polypropylene and the rubber-like substance  522  is a colored substance such as a colored thermoplastic elastomer. Alternatively, the rubber-like substance  522  could be clear and does not need to be colored. Each window  524  has a bottom side  526  that is located within an inner surface of the neck  514 . In the embodiments shown in FIGS.  5 D- 5 E, each window  524  has a flat bottom side  526  that is flush with a top of the rim  506  of the well  504 . In another embodiment shown in FIG. 5F, each window  524  has a flat bottom side  526  that is located within the well  504 . In yet another embodiment shown in FIG. 5G, each window  524  has a flat bottom side  526  that is located above the well  504 . In still yet another embodiment shown in FIG. 5H, each window  524  has a concaved bottom side  526 . Of course, each window  524  may have a bottom side  524  with other shapes and locations other than those described above with respect to FIGS.  5 D- 5 H.  
     [0033] To grow one or more protein crystals using the lid  500  and microplate  502 , the researcher pipets a small amount (e.g., 1.0 millimeter) of a reagent solution  530  into each of the wells  504  (see, FIGS.  5 D- 5 H). One or more drops  532  (only one shown) including a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter) that can be taken from the wells  504  are pipetted onto the bottom sides  526  of the necks  514 . Thereafter, the researcher inverts the lid  500  so the drops  532  hang down from the lid  500  and then positions, pushes-down and secures the lid  500  onto the microplate  502 . In this position, the drops  532  via a vapor diffusion process turn into protein crystals by interacting with the higher concentrated reagent solution  530  located within the wells  504  of the microplate  502 . Instead of having the researcher handle the lid  500 , the frame  512  of the lid  500  can have a footprint which makes it capable of being handled by a robotic handling system (not shown).  
     [0034] Referring to FIGS.  6 A- 6 H, there are illustrated different views of a third embodiment of a lid  600  designed to cover the individual wells  604  of a microplate  602 . The microplate  602  includes an array of wells  604  each of which has a rim  606 , sidewalls  608  and a bottom  610 . The wells  604  are generally arranged in a matrix of mutually perpendicular rows and columns. For example, the microplate  602  can include a matrix of wells  604  having dimensions of 4×6 (24 wells), 8×12 (96 wells) and 16×24 (384 wells). The microplate  602  shown includes an array of ninety-six wells  604 .  
     [0035] The lid  600  (e.g., protein crystallography hanging drop lid  600 ) is sized and configured to individually cover each of the wells  604  in the microplate  602 . In particular, the lid  600  includes a frame  612  having formed therein a series of downwardly protruding necks  614  (see cross-sectional side view in FIG. 6B and partial bottom view in FIG. 6C). Each neck  614  is designed to fit and seal against one of the wells  604  in the microplate  602 . In one embodiment shown in FIG. 6D, each neck  614  is designed to fit and seal against an inside edge of the rim  606  extending from one of the wells  604  in the microplate  602  (see also FIG. 6B). In another embodiment shown in FIG. 6E, each neck  614  is designed to fit and seal against an outside edge of the rim  606  extending from one of the wells  604  in the microplate  602 .  
     [0036] To ensure a relatively tight seal between each neck  614  and each well  604 , all or a portion of the bottom side and a portion of the top side of the frame  612  and an outer surface of each neck  614  are covered with a rubber-like substance  622  (shown as shaded area). In addition, as can be best seen in FIGS.  6 D- 6 H, the rubber-like substance  622  is located within portions of the frame  612  itself and on part of the top side of the frame  612  (compare to lids  400  and  500  in FIGS. 4B and 5B). The location of the rubber-like substance  622  within the frame  612  itself can help the rubber-like substance to adhere to the frame  612  better than if it was just located on the top side and/or bottom side of the frame  612 . In the preferred embodiment, the rubber-like substance  622  is a thermoplastic elastomer. However, any rubber-like substance  622  can be used that has gasket-like characteristics with the appropriate flexure and friction properties that can ensure a consistently tight seal around the wells  604 . The relatively tight seal between each neck  614  and each well  604 , should be maintained regardless of the different dimensional tolerances of the wells  604 , the hand assembly of the lid  600  to the microplate  602  and the movement of the lid  600  and microplate  602 .  
     [0037] Each neck  614  has a transparent window  624  which is part of the frame  612  that enables a user to see inside each well  604  when the lid  600  is attached to the microplate  602 . The window  624  is transparent because the frame  612  is preferably made from a clear substance such as polystyrene, cyclic olefin or polypropylene and the rubber-like substance  622  is a colored substance such as a colored thermoplastic elastomer. Alternatively, the rubber-like substance  622  could be clear and does not need to be colored. Each window  624  has a bottom side  626  that is located within an inner surface of the neck  614 . In the embodiments shown in FIGS.  6 D- 6 E, each window  624  has a flat bottom side  626  that is flush with a top of the rim  606  of the well  604 . In another embodiment shown in FIG. 6F, each window  624  has a flat bottom side  626  that is located within the well  604 . In yet another embodiment shown in FIG. 6G, each window  624  has a flat bottom side  626  that is located above the well  604 . In still yet another embodiment shown in FIG. 6H, each window  624  has a concaved bottom side  626 . Of course, each window  624  may have a bottom side  624  with other shapes and locations other than those described above with respect to FIGS.  6 D- 6 H.  
     [0038] To grow one or more protein crystals using the lid  600  and microplate  602 , the researcher pipets a small amount (e.g., 1.0 millimeter) of a reagent solution  630  into each of the wells  604  (see, FIGS.  6 D- 6 H). One or more drops  632  (only one shown) including a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter) that can be taken from the wells  604  are pipetted onto the bottom sides  626  of the necks  614 . Thereafter, the researcher inverts the lid  600  so the drops  630  are hanging down from the lid  600  and then positions, pushes-down and secures the lid  600  onto the microplate  602 . In this position, the drops  632  via a vapor diffusion process turn into protein crystals by interacting with the higher concentrated reagent solution  630  located within the wells  604  of the microplate  602 . Instead of having the researcher handle the lid  600 , the frame  612  of the lid  600  can have a footprint which makes it capable of being handled by a robotic handling system (not shown).  
     [0039] Referring to FIGS.  7 A- 7 C, there are illustrated partial cross-sectional side views of lid  600  having incorporated therein a microfluidic channel  702   a ,  702   b  and  702   c  that is associated with each neck  614 . The microfluidic channel  702   a ,  702   b  and  702   c  enables the researcher to load a drop  632  from the top side of the lid  600  and have the drop  632  slide onto the bottom side  626  of the neck  614 . In particular, the drop  632  can move via microfluidic transfer or capillary transfer through the microfluidic channel  702   a ,  702   b  and  702   c  onto the bottom side  626  of the neck  614 . Alternatively, the research can use a liquid handling device such as a pipet to force fluid through the micro-channel  702   a ,  702   b  and  702   c  to load the drop  632  on the bottom side  626  and the neck  614 . After loading all the drops  632  so that they are hanging from all the necks  614 , the researcher can seal the lid  600  using clear tape (not shown) to prevent the evaporation of the drops  632  and the reagent solutions  630  in the wells  604 . In other words, the researcher can connect the lid  600  to the microplate  602  and then load the drops  632  from the top side of the lid  600  onto the bottom sides  626  of the necks  614 . It should be noted that lids  400  and  500  in addition to lid  600  can also incorporate the microfluidic channels  702   a ,  702   b  and  704   c.    
     [0040] Referring to FIG. 8, there is a flowchart illustrating the steps of a preferred method  800  for using the lid  400 ,  500  and  600  and the microplate  402 ,  502  and  602 . The lid  400 ,  500  and  600  of the present invention is generally described as being coupled with the microplate  402 ,  502  and  602  to form a protein crystallography system. The protein crystallography system can be used to grow protein crystals using a hanging drop vapor diffusion crystallization process.  
     [0041] Beginning at step  802 , the lid  400 ,  500  and  600  is prepped by depositing one or more drops  432 ,  532  and  632  onto the bottom side  426 ,  526  and  626  of each neck  414 ,  514  and  614 . As described above, each drop  432 ,  532  and  632  includes a small amount of a protein sample (e.g., 1.0 microliter) and a small amount of a reagent solution (1.0 microliter).  
     [0042] At step  804 , the microplate  402 ,  502  and  602  is prepped by depositing the reagent solution  430 ,  530  and  630  into one or more wells  404 ,  504  and  604 . As described above, the researcher would pipet a small amount (e.g., 1.0 millimeter) of the reagent solution  430 ,  530  and  630  into each well  404 ,  504  and  604  of the microplate  402 ,  502  and  602 . The reagent solution  430 ,  530  and  630  located in each well  404 ,  504  and  604  would have a higher concentration than the reagent solution in the drops  432 ,  532  and  632 . It should be understood that the order of steps  802  and  804  can be reversed to enable the researcher to take a small amount of the reagent solution  430 ,  530  and  630  from the wells  404 ,  504  and  604  to form each drop  432 ,  532  and  632 .  
     [0043] At step  806 , the lid  400 ,  500  and  600  is placed over and pushed onto the microplate  402 ,  502  and  602  such that each neck  414 ,  514  and  614  fits and seals against the rim  406 ,  506  and  606  of each well  404 ,  504  and  604 . In this position, the drops  432 ,  532  and  632  on the bottom side  426 ,  526  and  626  of the necks  414 ,  514  and  614  can interact via a vapor diffusion process with the reagent solution  430 ,  530  and  630  within each well  404 ,  504  and  604  which enables the formation of protein crystals on the bottom sides  426 ,  526  and  626  of necks  414 ,  514  and  614 . Of course, if microfluidic channels  702   a ,  702   b  and  702   c  are incorporated into the lid  400 ,  500  and  600 . Then the drops  432 ,  532  and  632  could be deposited onto the bottom sides  426 ,  526  and  626  of the necks  414 ,  514  and  614  after the lid  400 ,  500  and  600  is attached to the microplate  402 ,  502  and  602 .  
     [0044] Referring to FIG. 9, there is a flowchart illustrating the steps of a preferred method  900  for making the lid  400 ,  500  and  600 . Beginning at step  902 , a first molten plastic material is injected into a first mold cavity that includes sections shaped to form the frame  412 ,  512  and  612  of the lid  400 ,  500  and  600 . Then at step  904 , the first plastic material is cooled to resemble the frame  412 ,  512  and  612 . Preferably, the first plastic material is a clear cyclic-olefin, polystyrene or polypropylene.  
     [0045] At step  906 , a second molten plastic material  422 ,  522  and  622  (e.g., rubber-like substance  422 ,  522  and  622 ) is injected into a second mold cavity that includes sections shaped to contain the frame  412 ,  512  and  612  and to enable the second plastic material  422 ,  522  and  622  to cover at least a portion of the outer surface,  520  and  620  of each neck  414 ,  514  and  614  formed in the frame  412 ,  512  and  612  (see FIGS. 4B, 5B and  6 B). Finally at step  908 , the second plastic material (e.g., rubber-like substance  422 ,  522  and  622 ) that was added to the frame  412 ,  512  and  612  is cooled to form the lid  400 ,  500  and  600 . Preferably, the second plastic material is a colored thermoplastic elastomer. Alternatively, the second plastic material could be clear thermoplastic elastomer.  
     [0046] The preferred method  900  can utilize at least two different molding processes to fabricate the two-component lid  400 ,  500  and  600 . One of these molding processes is generally known as overmolding or insert molding wherein the frame  412 ,  512  and  612  would be molded on a separate machine and then placed into a second machine whose mold cavity accepts the frame  412 ,  512  and  612  and also has a detail for the addition of the rubber-like substance  422 ,  522  and  622 . Another one of these molding processes is generally known as two-shot molding which uses one machine that first molds the frame  412 ,  512  and  612  and then moves the mold cavity to reveal a second stage mold which enables the rubber-like substance  422 ,  522  and  622  to be injected over the frame  412 ,  512  and  612  which never leaves the machine. Either of these processes or similar processes can be used to fabricate the lid  400 ,  500  and  600 .  
     [0047] Although several embodiments of the present invention has been illustrated in the accompanying Drawings and described in the foregoing Detailed Description, it should be understood that the invention is not limited to the embodiments disclosed, but is capable of numerous rearrangements, modifications and substitutions without departing from the spirit of the invention as set forth and defined by the following claims.