WO2000045953A1

WO2000045953A1 - Device for selectively regulating the temperature of individual containers

Info

Publication number: WO2000045953A1
Application number: PCT/EP2000/000899
Authority: WO
Inventors: Hans Lange; Thomas Kaplan; Gerhard Bienhaus; Jörg BÖTTGE
Original assignee: Bilatec Gesellschaft Zur Entwicklung Biotechnologischer Systeme Mbh
Priority date: 1999-02-05
Filing date: 2000-02-04
Publication date: 2000-08-10
Also published as: DE19904716A1; JP2002536155A; EP1148948A1

Abstract

The invention relates to a device (40) for selectively regulating the temperature of at least one container (3) of an aggregate (30) which represents a plurality of such containers (3). Said device comprises a receiving plate (1) for the container aggregate (30) which serves to maintain the container aggregate (30) at a predetermined temperature. The receiving plate (1) is provided with at least one opening through which one temperature-regulating element (2) is contacted with a container (3) whose temperature is to be regulated for the purpose of heat exchange. The container aggregate (30) and the one temperature-regulating element (2) can be moved relative towards one another. At least one additional cooling zone can be provided which dissipates the heat given off by the temperature-regulating element (2) that has not been used to regulate the temperature of the container (3).

Description

Device for the selective tempering of individual containers

description

The subject of the present patent application is a device for the selective tempering of individual containers, in particular for use in molecular biology and in screening, especially in high-throughput screening (HTS).

Pharmaceutical research was revolutionized by the possibilities of combinatorial chemistry because many different substances with pharmacological potential can be produced very efficiently using these methods. The starting point of pharmaceutical research today is a substance library in which a large number of chemical compounds, for example natural products, but also artificial peptides and nucleic acids are kept. In so-called high-throughput screening, a large number of these substances from the substance library are examined for their pharmacological effects using biological test methods in a partially automated process. Among other things, also molecular biological techniques applied. The entire process generally runs using pipetting and handling robots. More recent techniques using miniaturized systems in this environment are described in WO 98/00231. Special containers for the HTS are described in DE 298 05 61 3 and WO 97/1 5394.

The containers in which the substances from the substance library are stored are generally composite containers, for example in the 96, 384 or 1 536 microtitration plate format. A composite container in 96-well microtitration plate format is described, for example, in US 4, 1 54.795. There are various systems, including so-called cluster racks, which are composite devices for holding a large number of tubes. However, other formats are also conceivable, such as, for example, reaction vessels with a conical one Form as they are used for example for the amplification of nucleic acids. As a rule, these containers are sealed by sealing. Recent developments here show possibilities to close such containers with individual lids. For example, DE 44 1 2 286 describes machine-accessible lids that can also be used in the HTS environment.

Usually, very temperature-sensitive substances are kept in the substance libraries that are stored at low temperatures, e.g. -20 ° C or -70 ° C must be stored. Repeated thawing and refreezing damage such substances and must therefore be avoided.

There are now different strategies for how the substance library is used in HTS:

1 . Several "daughter plates" are pipetted from a so-called "mother plate". The "daughter plates" are then fed to the corresponding biological assay in the HTS.

2. Only predetermined individual connections, i.e. Individual containers, the substance library, are used to compile a selection of substances in a "daughter board", which is then fed into the HTS process.

The latter process in particular makes it necessary to thaw individual tubes or containers of a composite, to remove some of the substances and to freeze them again after sealing.

In order not to have to thaw and freeze all the containers on a plate unnecessarily often, devices are already known in the prior art with the aid of which individual reaction vessels of a composite are selective can be tempered, for example thawed, while the remaining containers remain frozen. These devices thus contribute to the longer shelf life of the sometimes valuable substances in a composite container and enable a longer usability of substance libraries.

For example, the older, but later published WO 99/1 6549 discloses a receiving plate for a group of 96 individual containers. Each of the individual receptacles of the receiving plate is equipped with its own heating device, which allows the individual container accommodated in this receptacle to be tempered, in particular thawed. The known temperature control device therefore has an extremely complex structure and is therefore correspondingly expensive to manufacture and sell. It is easy to see that the effort made here for 96 individual containers when using microtitration plates in the 384 or 1 536 format assumes economically unacceptable dimensions.

DE 1 900 279 discloses a temperature control device with a plurality of temperature control elements which can be brought into heat exchange contact together with a corresponding plurality of containers of a group of containers. For this purpose, the temperature control elements can be displaced in the vertical direction, while the container assembly can be displaced relative to the temperature control elements in the horizontal plane. It is disadvantageous, however, that the heat emanating from the temperature-regulating elements can spread unhindered in the group of containers, and there is therefore the risk that, when a row of containers is selectively thawed, at least the immediately adjacent rows of containers are also undesirably thawed.

Furthermore, reference is made to DE 34 41 1 79. However, the temperature control device for microtitration plates described there does not include any selective thawing unit. Rather, the important thing here is to temper the reaction vessels of the composite as evenly as possible. The same applies to DE 39 41 1 68 and DE 39 38 565.

Finally, for further prior art, reference is made to DE 1 573 1 93, DE 33 07 572, DE 40 22 792, DE 1 96 46 1 1 5, DD 239 473, DD 276 547, and US 4 , 1 1 6,777, US 4,950,608, US 5,849,208, and WO 98/1 5969.

In contrast, it is an object of the invention to provide a device which, with a simple and inexpensive construction, allows individual containers of a group of containers to be selectively tempered, and at the same time ensures that adjacent containers which are not to be tempered would be significantly influenced thereby, and in particular ensures that when Thawing individual containers The other containers in the group of containers are also not at risk of being defrosted.

According to the invention, this object is achieved by a device having the features of claim 1. In this device, the containers of the group of containers not to be tempered can be kept at a temperature by means of the receiving plate, which ensures that they remain frozen. The containers to be tempered by the heat exchange contact with the temperature control element, however, are arranged in the area of an opening in the mounting plate and are therefore not cooled by the latter. The relative movements of the container assembly and temperature control element (s) can be brought about in a simple manner by a robot. Such robots are in any case present and fall in the form of pipetting or handling robots in modern systems, for example for carrying out molecular biological tests on samples accommodated in the containers of a group of containers therefore not important in determining the cost of the device according to the invention.

A particular aspect of the present invention lies in the provision of at least one additional cooling zone which dissipates the heat given off by the temperature control element and not used for temperature control of the container. This additional cooling zone further increases the security against an undesired heat influence of containers of the group of containers that are not to be tempered. Independent protection is therefore also sought for this aspect.

The hybridization of single-stranded nucleic acids may be mentioned as a further application of a device according to the invention for the selective tempering of individual containers of a group of containers. In order to achieve the highest possible stringency of the hybridization, used a process that requires the temperature of the reaction vessel to a specific hybridization temperature - specific for each sequence. If several containers with different nucleic acid sequences are now to be hybridized in one work step, a device according to the invention is of considerable advantage, since several vessels can then be kept in parallel at different hybridization temperatures.

The invention will be explained in more detail below using exemplary embodiments with reference to the accompanying drawing. It shows:

Figure 1 is a rough schematic perspective view of a temperature control device according to the invention.

FIG. 2 shows a top view of the device according to FIG. 1;

3 shows a detailed view of a tempering mandrel; 4 shows a roughly schematic side view of the temperature control device according to FIGS. 1 and 2;

5a, 5b representations of a composite plate of a plurality of individual containers, with FIG. 5a a perspective

Bottom view and Fig. 5b shows a dimensioned side sectional view; and

Fig. 6-8 roughly schematic representations of further embodiments of temperature control devices according to the invention.

The arrangement according to the invention is based on two elements, namely on the one hand the special containers for storing and possibly treating the substances in the substance library and on the other hand the device for selective tempering.

5a shows a composite container 30 formed in the 96-microtitration plate format. The composite container 30 comprises a plate 11, on the underside of which a plurality of individual containers 3 are molded, for example by injection molding. It is characteristic that the individual reaction containers 3 are arranged freely below the composite plate 11 and without connecting webs. At the top of the composite panel 1 1, the openings 1 3 of the containers 3 are provided (see Fig. 5b). These can be closed with individual lids, such as those e.g. are known from DE 44 1 2 286. However, other closure options are known to the person skilled in the art.

In addition to the arrangement shown in FIGS. 5a and 5b, other embodiments are also possible. For example, the individual containers 3 do not necessarily have to be combined to form a permanent network 30. Alternatively, it is also possible to use a plurality of "loose" containers which are formed separately from one another a special perforated plate provided for this purpose (also referred to in technical terms as "rack") at least temporarily or temporarily to form a composite.

According to the invention, it is only necessary that the reaction containers 3 are freely accessible in their lower part. Thus, the distance A between the individual reaction containers 3 can be between approximately 1 mm and approximately 10 mm, preferably between approximately 2 mm and approximately 5 mm. 5b, the distance A has a value of 3.25 mm.

Furthermore, the distance B from the edge of the openings 13 to the bottom of the container 3 can be between approximately 1 mm and approximately 100 mm, preferably between approximately 5 mm and approximately 35 mm. 5b, the distance B has a value of 22.6 mm. The length of the zone 14 freely hanging underneath the plate 11 (see FIG. 1) in the exemplary embodiment according to FIG. 5 is 1 8 mm.

The shape of the containers 3 tapers at least over part of the distance from the opening 13 to the floor, preferably conically, so that there is sufficient space for the positioning of a tempering or Thawing mandrel 2 (see for example Fig. 2) is available. For rapid thawing or freezing, it is also advantageous if the individual containers in the free-standing zones 14 below the composite plate 11 are thin-walled in order to promote rapid heat exchange. Wall thicknesses of between approximately 0.2 mm and approximately 0.5 mm, in particular between approximately 0.2 mm and approximately 0.3 mm, are particularly advantageous.

In order to keep the exchange of heat via the plate 11 as low as possible, its thickness should be as low as possible. However, this is detrimental to the stability of the container assembly 30. The The thickness of the composite panel 11 should therefore be between approximately 0.5 mm and approximately 5 mm, preferably between approximately 2 mm and approximately 4 mm. 5b, the plate 11 has a thickness of approximately 4 mm.

The composite containers 3 can, for example, be made of thermally deformable plastic, such as polyethylene, polypropylene, polystyrene, polycarbonate, polyurethane or the like. They can be manufactured simply and inexpensively using injection molding processes.

For the selective tempering of one or more individual containers 3 of the composite container 30, the tempering device 40 shown in more detail in FIGS. 1 to 4 is used according to the invention. 1, the device 40 comprises a mounting plate 1, which is preferably made of a heat-conducting material, for example metal, preferably aluminum, brass or copper. The plate 1 can be cooled with the aid of cooling devices familiar to those skilled in the art, for example Peltier elements, compressor cooling units in conjunction with suitable heat transfer elements or the like. The purpose of this cooling is to keep the substances accommodated in the composite container 30 frozen after removal from the freezer.

In the plate 1 there are recesses 4 which serve as receptacles for the containers 3 and whose shape is therefore chosen in accordance with the outer contour of the containers 3. In the embodiment shown, the depressions 4 are designed as longitudinal grooves, so that the containers 3 can be displaced in the direction of the double arrow y, ie in the Y direction. Approximately in the middle of the displacement distance of the plate 1, each longitudinal groove is provided with at least one opening 5. A temperature control element, for example designed as a temperature control or thawing mandrel 2, can be inserted into each of the openings 5. The task of this tempering or thawing mandrel 2 is to temper or thaw the container 3 arranged in the area of the respective opening 5. For this purpose, the temperature control element 2 can be brought into physical heat exchange contact with the surface of the respective container 3 by a vertical movement in the direction of the arrow z, ie in the Z direction. To use the temperature control device according to the invention for multiple hybridization at different temperatures, several temperature control elements 2 are inserted into several openings 5.

According to FIG. 2, the openings 5 are surrounded by cooling zones 1 6, of which only one is shown in FIG. 2. These cooling zones serve to dissipate the heat emitted by the tempering or thawing mandrel 2, which is not used for tempering or thawing the container 3, as effectively as possible, so as to be able to prevent an undesirable influence of this heat on neighboring containers 3. The cooling zones 16 can, for example, comprise ring elements made of a thermally highly conductive material, which are thermally coupled to an external cooling device. The ring elements can, for example, be full rings made of copper or the like, or else ring tubes through which a cooling fluid flows. If several such additional cooling zones are provided, at least some of these cooling zones can be thermally coupled to a common cooling device. The temperature difference between the cooling zones 16 and the coolable receptacle 1 is advantageously between approximately 5 ° C. and approximately 50 ° C., preferably between approximately 20 ° C. and approximately 30 ° C.

In the embodiment according to FIG. 2, each of the longitudinal grooves 4 has a single opening 5. In addition, a linear movement unit 7 is provided with a movable rake 6, by means of which the composite container 30 accommodated in the plate 1 can be moved in the Y direction . This ensures that each Container 3 of the composite container 30 can be pushed over a tempering or thawing position 5. Furthermore, the tempering or thawing mandrel 2 is designed to be displaceable in the X direction. By moving the composite container 30 in the Y direction and the dome 2 first in the X and then in the Z direction, any container 3 of the composite container 30 can thus be controlled and thus tempered or thawed.

In principle, however, it is also possible for the receiving plate 1 to have a number of receptacles corresponding to the number of containers 3 of the container assembly 30, each of which is provided with an opening 5, and for the temperature control or thawing element 2 to be movable in all three spatial directions is, ie both in the X, Y and Z directions.

3 shows the structure of the temperature control or thawing element 2 used according to the invention. It contains an insulating layer 8, which ensures that the heat transfer between the tempering or thawing mandrel 2 and the environment, for example the metal plate 1, is minimal. As a result, the radiation or emission of heat not used for tempering the vessel 3 in question can be kept low. The interior of the temperature control or thawing mandrel 2 is preferably made of ceramic material 8 with an electrically heatable coil 9. In principle, microwave or infrared heaters could also be used. However, heating coils embedded in a suitable material, such as ceramic, have the advantage that their use, particularly in reaction mixtures with biologically active substances, does not damage these substances by radiation effects or the like.

FIG. 4 shows another inventive detail in a special embodiment: If the tempering or thawing mandrel 2 moves upwards, it must be against the one to be thawed in order to achieve good heat transfer Container are pressed. However, there is a risk that the container assembly 30 will thereby be lifted out of the coolable receptacle 1. To prevent this, when the temperature control element 2 is raised, a counter-holding device, for example a counter-holding plate 15, is automatically shut down, which presses the container assembly 30 into the receiving grooves 4 of the coolable plate 1. When using a device for automatically removing or fitting a lid assigned to the individual container to be tempered, the counter-holding device can also be formed by a gripper arm of this lid handling device. This further simplifies the construction of the device according to the invention, since no separate counter-holding device needs to be provided.

Surprisingly, it was found that the structure explained above enables the sample containers 3 to be heated up very quickly without the risk that their surroundings, i.e. the containers 3 surrounding them would be thawed by the selective heating process. This arrangement allows, by appropriate temperature setting and control of the temperature control elements 2, the selective temperature control of one or more containers 3 to a temperature which is between approximately 20 ° C. and approximately 70 ° C. above the temperature of the remaining containers.

In addition to the device described above, in which this assembly 30 and a tempering element 2 had to be positioned in a plurality of linear relative movements relative to one another in order to temper individual containers 3 of a group of containers 30, there are also differently designed devices for selectively tempering individual containers 3. In the figures 6 to 8, three such devices are shown roughly schematically by way of example only. The basic principle of these devices corresponds to the temperature control device known from the older, but later published WO 99/1 6549, can according to the invention however, all be equipped with additional cooling zones 1 6, as explained above.

In the embodiment according to FIG. 6, 4 Peltier elements 18 are attached to each receptacle in such a way that a good thermal contact surface is created in the receptacle 4. It is known to the person skilled in the art that Peltier elements consist of a cooling side 19 and a heating side 20 and these can be exchanged accordingly by electrical polarity reversal. A corresponding mounting plate 1, as shown in FIG. 6, now has the possibility of holding reaction containers in a composite and cooling them by switching the Peltier elements accordingly. Appropriate controls can be used to selectively temper individual containers by reversing the polarity of the Peltier elements. In order to be able to set an exact temperature control accordingly, it can be advantageous to add or remove heat accordingly via a further plate 17 in FIG. 6 in order to simplify and specify the control. For this purpose, according to the invention, a temperature-adjustable plate 17 as in FIG. 6 (here, for example, on the floor) can be advantageous. In the case of the thawing application, the plate 1 7 should be cooled to dissipate the heat of heating.

Another embodiment according to the invention is shown in FIG. 7. Here individual receptacles 1 with depressions 4 for receiving containers 3 on the bottom are coupled to a Peltier element 1 8. The individual recordings are separated from one another by a heat insulation layer 28. A corresponding air duct 21 is attached to the floor for dissipating the heat and ensures the cooling of the Peltier elements 1 8 by means of a fan 1 2. Comparable to FIG. 6, the polarity reversal of the Peltier elements 18 can be used to achieve a selective temperature control or heating.

8, cooling or heating liquid is supplied to the surroundings of the receptacles 4 for the containers 3. The corresponding coolable Receptacles 1 are individually separated from one another by an insulating layer 28 and each have bores 22 which can be charged with cooling or heating liquid. These can be corresponding cooling coils or, if the coolable receiving plate is made of metallic materials, corresponding bores with connections. The individual connections can all be controlled with a corresponding valve system so that individual receptacles can be selectively cooled or heated.

The devices according to FIGS. 6 to 8 are particularly suitable for nucleic acid hybridization in several containers with different sequences and thus hybridization temperatures. It should once again be expressly emphasized that these three embodiments are possibilities for an inventive implementation.

Claims

Expectations

1 . Device (40) for the selective tempering of at least one container (3) of a composite (30) having a plurality of such containers (3), the device comprising a receiving plate (1) for the container assembly (30), which serves to connect the container assembly (30) 30) to be kept at a predetermined temperature, the receiving plate (1) having at least one opening

(5), by means of which at least one temperature control element (2) can be brought into heat exchange contact with a container (3) to be temperature-controlled, and the container assembly (30) and the at least one temperature control element (2) can be moved relative to one another.

2. Device according to claim 1, characterized in that the receiving plate (1) comprises a plurality of longitudinal grooves (4), in each of which a plurality of containers (3) of the container assembly (30) can be received.

3. Device according to claim 2, characterized in that each of the longitudinal grooves (4), preferably in the area of approximately their longitudinal center, has at least one opening (5).

4. The device according to claim 3, characterized in that a drive device (Y) is provided for positioning the container assembly (30) relative to the at least one opening (5), preferably by moving the container assembly (30) in the longitudinal grooves.

5. The device according to claim 3 or 4, characterized in that a drive device (X, Z) is provided for positioning the at least one temperature control element (2) relative to the at least one opening, preferably by moving the temperature control element in a first to the longitudinal grooves ( 4) orthogonal, but essentially parallel to the plane of the mounting plate (1) and a second direction (Z) running essentially orthogonal to the plane of the mounting plate (1).

6. Device according to one of the preceding claims, characterized in that a counter-holding device (1 5), for example a counter-holding plate (1 5), is provided, which presses the container assembly (30) against the receiving plate (1).

7. Device according to one of the preceding claims, characterized in that at least one additional cooling zone (1 6) is provided, which dissipates the heat given off by the temperature control element (2) and not used for temperature control of the container (3).

8. Device (40) for the selective tempering of at least one container (3) of a composite (30) having a plurality of such containers (3) by means of at least one tempering element (2), if desired according to one of the preceding

Claims, wherein the device comprises a receptacle plate (1) for the container assembly (30), which serves to keep the container assembly (30) at a predetermined temperature, and wherein at least one additional cooling zone (1 6) is provided, which the Tempering element (2) dissipated and not used for tempering the container (3) dissipates heat.

. Apparatus according to claim 7 or 8, characterized in that the additional cooling zone (1 6) comprises a ring element made of thermally highly conductive material, which is thermally coupled to a cooling device.

0. Device according to one of claims 7 to 9, characterized in that when a plurality of additional cooling zones (1 6) are provided, at least some of these cooling zones (1 6) are thermally coupled to a common cooling device.

1 . Device according to one of claims 7 to 10, characterized in that the temperature difference between the receiving plate (1) and the additional cooling zone (1 6) is between approximately 5 ° C and approximately 50 ° C, preferably between approximately 20 ° C and approximately 30 ° C.

2. Device according to one of the preceding claims, characterized in that the temperature control element (2) comprises at least one Peltier element.

3. Device according to one of the preceding claims, characterized in that the temperature control element (2) comprises at least one heating coil.

4. Device according to one of the preceding claims, characterized in that the temperature control element (2) is surrounded by a heat insulation layer (8).

5. Device according to one of the preceding claims, characterized in that a plurality of temperature control elements

(2) is provided, which can preferably be tempered independently of one another.

1 6. Device according to one of the preceding claims, characterized in that the container assembly (30) comprises a composite plate (1 1) on which the plurality of containers (3) is integrally formed.

1 7. Device according to one of the preceding claims, characterized in that the distance (A) between adjacent containers (3) is between approximately 1 mm and approximately 10 mm, preferably between approximately 2 mm and approximately 5 mm.

1 8. Device according to one of the preceding claims, characterized in that the distance (B) from the composite plate (1) to the bottom of the containers (3) is between about 1 mm and about 1 00 mm, preferably between about 5 mm and about 35 mm.

1 9. Device according to one of the preceding claims, characterized in that the wall thickness of the containers (3) is between about 0.2 mm and about 0.5 mm, preferably between about 0.2 mm and about 0.3 mm.

20. Device according to one of the preceding claims, characterized in that the thickness of the composite plate (1 1) is between approximately 0.5 mm and approximately 5 mm, preferably between approximately 2 mm and approximately 4 mm.

21. Use of a device according to one of the preceding claims for screening, preferably high-throughput screening, for example in molecular biology, such as in nucleic acid hybridization.