1. Field of the Invention
The present invention relates to devices and methods for dosing small amounts of liquid, and in particular to devices and methods which are suitable for simultaneously and accurately discharging small or extremely small amounts of dosed liquid from a plurality of parallel channels.
2. Description of Prior Art
The accurate dosing of amounts of liquid is of essential importance e.g. in the fields of pharmaceutical and biotechnological research, e.g. in the field of genomics, high-throughput screening, combinatorial chemistry and the like. Such dosing is e.g. necessary for filling so-called well-plates with reagents. In order to realize such filling various devices and methods are known at present; the most common ones of these devices are air-cushion pipettes, valve-controlled piston-displacement pipettes, piezoelectric pipettes and needle pipettes. The devices referred to are typically of the single-channel type. Part of these devices can, however, also be arranged in a raster so that parallel channels are provided. The highest degree of parallelization achieved is, at present, 384 channels in the case of some commercially available devices used for dosing volumes above 0.5 μl.
When the above-mentioned principles and devices are used for filling well-plates, one or a plurality of the problems mentioned hereinbelow frequently occur(s). In most cases, it is impossible to discharge dosing volumes which are smaller than 500 nl. This applies especially to air-cushion pipettes. In addition, the accuracy of all commercially available devices is unsatisfactory in the lower dosing range, the error exceeding typically 10%. Furthermore, due to the structural design of the conventional devices, it is impossible to achieve a high level of integration, e.g. raster dimensions of less than 4.5 mm, by said devices so that serial processing has to be carried out in some cases. Only needle pipettes are suitable for achieving also raster dimensions which are as small as 2.25 mm. If dosage is not effected in the form of a free jet, as e.g. in the case of needle pipettes, displacements and cross-contaminations of the liquids to be dosed may occur.
Known microdosage devices are described in DE-A-19706513 and in DE-A-19802368. These known devices are based on a functional principle according to which a liquid to be dosed has applied thereto an acceleration by a displacer within a pressure chamber. The pressure chamber is in fluid connection with an outlet opening and with a fluid reservoir. It follows that, when the displacer these known devices is operated, a movement of the liquid through the outlet opening as well as back into the reservoir will take place.
DE-A-19913076 discloses a microdosage device by means of which a plurality of microdroplets can be applied to a substrate; in this microdosage device the whole dosing head is acted upon by an acceleration. Due to this acceleration of the whole dosing head, an inertia-dependent relative acceleration between the fluid contained and the dosing head is achieved, said relative acceleration being of such a nature that droplets are ejected from the respective nozzle openings.
Finally, WO 00/62932 discloses methods and devices for discharging extremely small, dosed amounts of liquids, the discharge amounts mentioned being in the range of from 0.1 nl to 100 μl. According to this publication a capillary is used, which is provided with a discharge opening and which has connected thereto at least one gas line via a junction point. Via the capillary, a gas blast is introduced in the gas line so that an amount of liquid contained in the capillary section between the junction point and the discharge opening will be discharged from the discharge opening in a dosed amount. This publication also mentions the possibility of producing a pipetting array making use of a plurality of dosage devices of the type described hereinbefore. Also in the dosage devices disclosed in this publication a return flow into the reservoir takes place and, in the most disadvantageous case, air bubbles may rise into the reservoir line and block it.
The above-mentioned disadvantages result in the fact that the time which is necessary to fill a well-plate is normally considered to be too long for the desired throughput. This results, on the one hand, in high costs and, on the other hand, partly also in difficulties as far as the analysis of the reaction products is concerned, if the reactions in the individual reservoirs of the well-plate start with a time shift.
In R. Zengerle, “Mikrosysteme—Chancen für die Dosiertechnik”, wägen+dosieren 1/1996, pp. 10-15, micropumps for microdroplet injectors are disclosed in the case of which the volume of a pump chamber can be varied by means of a diaphragm which is adapted to be driven by a piezo bending transducer. An inlet opening and an outlet opening of the pressure chamber are provided. A pumping effect can be achieved in response to actuation of the diaphragm, in that either the inlet opening and the outlet opening are provided with a non-retum valve or in that a buffer is provided adjacent the pressure chamber.
DE 19648694 C1 discloses a bidirectional dynamic micropump comprising a pump chamber as well as an inlet and an outlet for the pump chamber with different flow resistances. A diaphragm borders on the pump chamber, whereby the delivery direction of the micropump can be controlled by suitably shaping the control pulse for the diaphragm.
WO 97/15394 discloses a plate having a plurality of apertures which extend therethrough. The apertures have a large opening towards one surface of the plate and a small nozzle opening towards the opposite surface of said plate. By applying a pressure on the large opening, a jet of liquid can be ejected through the small nozzle opening.
Another dosage device is known from WO 99/36176, said dosage device comprising a liquid reservoir and a channel which is in fluid communication with the liquid reservoir. Openings are formed in opposed walls of the channel so that the liquid present between the openings can be discharged, in a dosed amount, by applying a pressure to one of the openings.
It is the object of the present invention to provide devices and methods for dosing small amounts of liquid, which permit a simple structural design of a microdosage device and which additionally permit an accurate, precise dosage of small amounts of liquid from a plurality of parallel channels.
According to a first aspect of the invention, this object is achieved by a microdosage device having the following features:
a media reservoir used for accommodating a liquid to be dosed;
a nozzle connected via a connecting channel to the media reservoir and adapted to be filled via said connecting channel with the liquid to be dosed; and
a drive unit for applying, when actuated, to a liquid contained in the media reservoir and in the nozzle a force of such a nature that a substantially identical pressure will be exerted on said liquid contained in the media reservoir and in the nozzle,
wherein flow resistances of the connecting channel and of the nozzle are dimensioned such that, in response to an actuation of the drive unit, a volumetric flow in the connecting channel will be small in comparison with a volumetric flow in the nozzle, said volumetric flow in the nozzle causing an ejection of the liquid to be dosed from an ejection opening of the nozzle.
According to a second aspect of the invention, the above object is achieved by a method for dosing small amounts of liquid, said method comprising the following steps:
filling at least one nozzle via a connecting channel with a liquid to be dosed from a media reservoir, said connecting channel establishing a fluid connection between said nozzle and said media reservoir;
applying to the liquid contained in the media reservoir and in the nozzle a force of such a nature that a substantially identical pressure will be applied to the liquid contained in the media reservoir and in the nozzle,
so that, due to the dimensioning of flow resistances of the connecting channel and of the nozzle, the volumetric flow in the connecting channel will be small in comparison with the volumetric flow of the liquid in the nozzle so as to eject an amount of the liquid to be dosed from an ejection opening of the nozzle.
The functional principle of the microdosage device according to the present invention and of the method for dosing small amounts of liquid according to the present invention is based on two points; firstly, that the reservoir and the nozzle and nozzle channel, respectively, have simultaneously applied thereto a force, and secondly that the reservoir and the nozzle are separated from one another to a sufficient extent by the connecting channel. The higher the fluidic resistance of the connecting channel in relation to the fluidic resistance of the nozzle channel is, the more effective such a separation will be. Due to the fluidic separation, the maximum amount dosed in the case of the present invention will be the volume contained in the nozzle; when this volume has been discharged, the dosing operation will stop automatically.
According to the present invention, the reservoir, the nozzle and the connecting channel are preferably formed in a dosing head; such a dosing head may preferably comprise a plurality of reservoirs, nozzles and connecting channels. In order to cause an ejection of droplets from the nozzle opening or the nozzle openings of said one or of said plurality of nozzles, a driving force is applied to the whole liquid contained in the dosing head in accordance with the present invention, i.e. both the reservoir and the nozzle are acted upon by this force. This is the reason for the fact that a return flow into the reservoir does not take place in the case of the present invention, another reason being that the pressure gradient along the connecting line is negligible.
In preferred embodiments of the present invention the reservoirs as well as the nozzles are arranged in a raster which corresponds to the format of a well-plate. Furthermore, the reservoirs and the nozzles may be arranged in different rasters so that a change of format between the format of the reservoirs and that of the receiving receptacle, which is normally a well-plate, is effected by a dosing process. The dosing head used in the microdosage device according to the present invention can be produced in a conventional, known manner making use of micro-mechanical methods; it can be produced from silicon or from plastic material, by way of example, making use of e.g. an injection moulding technology. In preferred embodiments the drive means consists of a pneumatic or a hydraulic drive unit comprising a pressure chamber that can rapidly be filled with a gas or a liquid as a buffer medium so as to apply the necessary force to the reservoirs and the nozzles.
According to the present invention, the reservoirs and the reservoir ends located opposite the ejection ends of the nozzles are preferably formed in a surface of the dosing head so that the whole first side of the dosing head and of the dosing head substrate, respectively, can be acted upon by the driving force; this application of force will have the effect that only the liquid contained in the nozzle, i.e. the nozzle channel and the nozzle opening, will be discharged and that the dosing process will stop automatically as soon as the liquid contained in these elements has been discharged. Making use of this principle, a spatial separation of the areas in which the nozzles and the reservoirs are arranged is no longer necessary, whereby the levels of integration that can be achieved will be substantially higher than those achieved by devices in which the driving force only acts on the rear nozzle areas, but not on the reservoirs.
It follows that the present invention provides devices and methods by means of which liquids can be discharged e.g. into a well-plate in a highly parallel mode. The microdosage device according to the present invention has therefore a simple structure but, nevertheless, it permits exact dosage even if the system realized is a highly integrated dosing system in which parallel dosage is to be carried out making use of e.g. 1536 parallel nozzles. According to the present invention, such precise dosage can be carried out without making use of active or passive valves, which are used in the prior art in some cases, since the reservoirs as well as the nozzles are acted upon by the force and since the connecting channel between these elements has a suitable structural design.
Hence, the present invention represents a substantial improvement of dosing technology in the nanolitre range, which permits a highly parallel and therefore a much faster dosage of reagents into well-plates. The present invention allows a high degree of parallelization and a high level of integration; it is, for example, possible to carry out 96, 384, 1536 or more dosing processes simultaneously in the case of raster dimensions of 9.0 mm, 4.5 mm, 2.25 mm or raster dimensions which are smaller than that. In addition, the present invention also permits an adaptation to formats outside of the standard for well-plates. The present invention also permits an extremely high accuracy of dosage, the dosing volume error being less than 5 nl in the case of typical dosing amounts of from 50 nl to 100 nl. Due to the contact-free discharge in a free jet, a displacement of media is additionally excluded. As has already been mentioned, re-formatting, e.g. from a 384 format to a 1536 format, can be carried out in parallel when the device is provided with a suitable structural design.
Furthermore, the present invention allows media to be stored in the dosing head so that it is no longer necessary to perform the operating step of transferring the media from the storage unit, which is nowadays typically a well-plate having 96 reservoirs, to the automatic dosage device, which consists nowadays of air-cushion pipettes or of similar devices arranged in parallel. Finally, the dosed volume is largely independent of the physical properties of the liquids used in the case of the device according to the present invention and the method according to the present invention. The present invention also permits the construction of a microdosage device in which the dosing head can be replaced easily so that the drive unit, which is normally more complicated and more expensive than the dosing head itself, can be used for a large number of different dosing heads. In this respect, it is, in particular, also of advantage that the entire first surface of the dosing head is acted upon by the force so that an adaptation is here not necessary, not even in the case of different arrangements of reservoirs and nozzles in different dosing heads.
Hence, the present invention is particularly suitable for discharging precise amounts of liquids into well-plates having standardized outer dimensions and comprising a large number of juxtaposed reservoirs. As has been stated hereinbefore, such well-plates comprise a large number of reservoirs, e.g. 96, 384, 1536 or more, the raster distances between the reservoirs being, accordingly, 9 mm, 4.5 mm, 2.25 mm, etc. Depending on the integration level, the volume of the reservoirs is approx. 100 μl, 20 μl, 4 μl, etc. In these reservoirs chemical and biochemical reactions are caused to take place and the reaction products are analyzed. The possibility of precisely filling well-plates with predetermined amounts of liquid is therefore an indispensable prerequisite for executing quantitative analyses making use of very small amount of liquids; the present invention specially offers this possibility in an advantageous manner.
In addition to precise filling of the well-plate, also a fast, preferably simultaneous dosage of reagents into all reservoirs is of interest, since, normally, a high number of reactions are caused to take place in a well-plate simultaneously. It will here be of advantage when the whole sequence of steps carried out for filling the well-plate and for analyzing the results can be automated, so that several hundred well-plates can be processed per day and so that a few thousand up to one hundred thousand reactions can be caused to take place. Due to the fact that the present invention has the property of allowing a strongly parallel and highly precise dosage of very small amounts of liquids, it is particularly suitable for this purpose, especially also for discharging a great variety of different liquids into the various reservoirs of a well-plate.
In the following, preferred embodiments of the present invention will be explained in detail making reference to the drawings enclosed, in which:
As can be seen in
As shown in the figure, the media reservoir 4 is arranged on a first side of the dosing head 2, i.e. it is formed in a first surface thereof, whereas the discharge of liquid takes place through the nozzle opening 22 on the opposite, second side of the dosing head. The unit which is here referred to as nozzle is defined by the nozzle channel 20 and the nozzle discharge opening 22 and represents a fluid connection between the first surface 24 and the second, opposite surface 26 of the dosing head 2. In the embodiment shown in
According to the present invention, the flow resistances of the connecting channel 10 and of the nozzle, i.e. of the nozzle channel 20 and of the nozzle opening 22, are dimensioned such that, when a liquid 8 in the media reservoir and in the nozzle is acted upon by a force of such a nature that a substantially identical pressure is applied to the liquid in the media reservoir and in the nozzle, a volumetric flow in the connecting channel 10 will be small in comparison with a volumetric flow in the nozzle 6.
This situation can be achieved when the flow resistance R2 of the connecting channel 10 is large in comparison with the flow resistance of the nozzle channel between the first side 24 and the second side 26 of the dosing head 2, i.e. when it is equal to R11+R12+R13. Furthermore a sufficiently good dosing quality can already be achieved in the embodiment shown in
As has already been mentioned hereinbefore, an exact dosage can be achieved by dimensioning the flow resistance R2 of the connecting channel such that it is much higher than the overall resistance of the nozzle channel. In cases in which the connecting channel 10 leads into the nozzle channel 20 in spaced relationship with the first side 24, so that a resistance R11 can be defined, a sufficiently good result will already be achieved when the condition R2>>R11 is fulfilled. With respect to the dimensioning of the resistances R11 and R2 it should be taken into account that the larger the difference between the resistances is the larger the bandwidth of the various liquids which, making use of a suitable dosage device, can be dosed with sufficient accuracy will be.
With respect to the dimensioning of the resistances R11 and R2, it should be taken into consideration that the dosed volume depends on the ratio of the two resistances. If R2/R11≈10 is chosen, the dosed volume will correspond to the liquid volume contained in the nozzle with a systematic deviation of 10% at the most. This deviation results from the fact that, due to the pressure drop across the flow resistance R11, the pressure prevailing at the location where the connecting channel leads into the nozzle will be lower than the pressure prevailing on the upper side, i.e. the first side, of the dosing head and in the reservoir, respectively. This has the effect that a pressure difference across the connecting channel will occur whose magnitude depends on the ratio of the flow resistances R2 and R11, said pressure difference inducing an additional volumetric flow in the connecting channel in the direction of the nozzle opening. This volumetric flow contributes to the dosed volume as well. The exact magnitude of this systematic deviation depends on the details of the concrete structural design of the connecting channel and of the nozzle channel. The deviation can be minimized by a skilful geometrical design of the channels especially at the junction point of these channels. The percentage to which the induced flow contributes to the overall flow through the nozzle and the nozzle opening, respectively, can, however, be estimated with the value R11/R2 towards the upper limit independently of these geometrical details. It follows that, due to the additional flow through the connecting channel, the dosing volume will at most increase to (1+R11/R2) times the volume of the nozzle channel.
Since the process described hereinbefore is, however, reproducible and since the ratio of the flow resistances does not depend on the media properties of the fluid, the accuracy and the functional efficiency of the dosage device will not be impaired by this increase. It follows that the exact ratio of the flow resistances is not of essential importance, as long as R11<R2. Summarizing, it can be stated that the ratio of the flow resistances causes a systematic error which can be compensated for when dosage devices are being produced. It does, however, not cause any statistical error which would influence the reproducibility of the dosage device. With respect to a simple and precise implementation of the dosage device, it may, in practice, be desirable to define the dosing volume as precisely as possible by the volume contained in the nozzle. In this case, it will advantageous to choose R11/R2 as small as possible and R2/R11 as large as possible, e.g. R2/R11>100. This will lead to excellent fluidic decoupling between the nozzle and the reservoir during a dosing operation and the dosed volume will correspond to the nozzle volume with a maximum deviation of 1%.
When the reservoir and the nozzle are acted upon by a force of such a nature that a substantially identical pressure is applied to the liquid in the reservoir and in the nozzle, this will guarantee in any case that no return flow through the connecting channel will take place according to the present invention. Such a return flow occurs in known dosage systems of the type described e.g. in the above-mentioned publications DE-A-19706513, DE-A-19802368 or WO 00/62932. A network model representative of the dosage devices shown in the above-mentioned publications is shown in
Making again reference to
It should here be pointed out that, in the embodiment shown in
When the nozzle has been filled completely, this filling being caused by capillary forces in preferred embodiments of the present invention, the dosing head 2 is connected to a drive unit, as schematically shown in
If the flow resistance of the connecting channel is sufficiently high in comparison with the flow resistance of the nozzle and in comparison with the resistance R11, the liquid in the connecting channel will essentially remain in a state of rest, whereas the liquid from the nozzle channel 20 will be ejected through the nozzle opening 22.
In such a dosing process the whole amount of liquid contained in the nozzle can be discharged through the nozzle opening 22, without any movement of the liquid contained in the connecting channel taking place. It follows that the dosed amount of liquid is precisely determined by the geometry of the nozzle. Dosage of the liquid will stop automatically when the nozzle has been emptied completely.
As has already been described, a fluid volume corresponding to the total volume of the nozzle 6 can be discharged from the nozzle opening 22 by actuating the drive unit. It is, however, also possible to eject through said nozzle opening only part of the amount of liquid contained in the nozzle and defined by the geometry of the nozzle, whereas the liquid contained in the connecting channel is not moved or only moved to an insignificant extent.
When the nozzle has been emptied fully or partly, the original condition can be reestablished, after deactivation of the drive unit, by two possibilities, alternatively. Firstly, the pressure chamber can be vented, e.g. by through the valve 16 shown in
If a vent means is not provided or if the drive unit uses mere switching valves, i.e. 2/2-way pneumatic valves, the overpressure can diminish due to a flow of gas through the nozzles when the pressure supply has been switched off.
When the overpressure in the pressure chamber 12 has been reduced to a sufficient extent, the connecting channel 10 and the nozzle 6, i.e. the nozzle channel 20 and the nozzle opening 22, will, due to capillary forces, be refilled from the media reservoir connected thereto, whereupon a renewed dosing process can be carried out.
In order to achieve a neat tearing of the liquid column discharged at the nozzle opening, it will be advantageous to produce in the microdosage device according to the present invention a sufficiently high pressure amplitude in the pressure chamber, whose variation with time should in addition advantageously take place within a very short period so as to achieve a high dynamic of pressure variation. Furthermore, it will advantageous to implement the dosing head and the drive unit such that the discharge of liquid will be finished within a short period of time, e.g. 10 milliseconds, whereas the fluid lines, which are used for refilling on the basis of capillary forces, are implemented such that this process will take much longer, e.g. 100 milliseconds. The two effects will therefore only overlap each other to an insignificant extent and the precision of the dosing volume will not be distorted by the capillary refilling process.
A microdosage device according to the present invention as well as the mode of operation of this device have now been described in general, and, in the following, embodiments and special further developments of this microdosage device will be described in more detail.
The embodiment shown in
Making use of these fast-switching valves 42, it will be possible to generate an overpressure with high dynamic in the pressure chamber 12; by means of this overpressure a driving force is transmitted to the liquids in the dosing head, i.e. it is simultaneously transmitted to the liquids contained in the media reservoirs 4 and in the nozzles 6.
It is evident that a micro-dosing head according to the present invention can comprise an almost arbitrary number of media reservoirs and nozzles;
Reference should here be made to the fact that, according to the present invention, it is not necessary that each nozzle has associated therewith a media reservoir, but that there is, essentially, a freedom of choice insofar as one or a plurality of media reservoirs can be provided which may have connected thereto one or a plurality of nozzles; in this case, a respective media reservoir can be connected to a nozzle via several connecting lines, a media reservoir can be connected to a plurality of nozzles via several connecting lines, and a nozzle can be connected to a plurality of media reservoirs via several connecting lines, as will be explained hereinbelow with reference to
Making reference to
The nozzle 6 shown in
Finally,
With respect to the structural design of the nozzle it should, finally, be stated that it is possible to design the nozzle discharge opening as well as the opening located opposite thereto and the nozzle channel, which connects these openings, not circularly but with an arbitrary shape. Deviating from that which is shown in
Various design possibilities for the connecting channel interconnecting the interior reservoirs and the nozzles are shown in
In the example shown, a circular reservoir 4 is connected to the nozzle 6 by a rectangular connecting channel 10a, the reservoir and the connecting channel having the same depth so as to guarantee that the reservoir will be emptied completely.
The connecting channel between the media reservoir and the nozzle may, in addition, have arbitrary cross-sections and need not necessarily be rectangular. Finally, the cross-section may become larger or narrower in the course of the channel, and it would also be possible to provide e.g. two or more connecting channels between the same reservoir and the same nozzle.
Such an embodiment is shown in
Irrespectively of whether an identical number of nozzles and reservoirs is provided, a change of format can be effected by the dosing head in any case. This means that the distance between the individual nozzles and the distance between the individual reservoirs can be different. One example of such an arrangement with broad distances between the reservoirs 4 and narrow distances between the nozzles 6 is shown in
As can be seen from the fragmentary, schematic, cross-sectional views shown in
Through the system liquid 46, the force is applied to the liquid to be dosed, whereupon the dosing process will take place.
As can clearly be seen from the above explanations, the dosed volume according to the present invention is determined by the volume of the nozzle, either only in the nozzle channel or in the nozzle channel and in the nozzle opening, and is essentially independent of the properties of the fluid. The present invention permits a precise discharge of e.g. approx. 50 nl in a single dosing process, provided that the channels and openings are designed appropriately.
Although preferred embodiments of the present invention have been explained in more detail hereinbefore, it is evident that further modifications and changes of these embodiments are possible. It is, for example, possible to use alternative drive units so as to exert a driving force on the liquid. In addition to the described application of a homogeneous, pneumatic or hydraulic pressure in the area of the first side of the dosing head, it is also possible to apply a negative pressure to the second side of the dosing head so as to suck the liquid from the nozzle opening. Still another alternative is a volume displacement of liquid on the first surface of the dosing head, one example of such a volume displacement being the embodiment which makes use of the system liquid 46 and which has been described making reference to
It need not be specially mentioned that the plurality of reservoirs of the dosing head used according to the present invention can be filled with identical or different liquids so that identical or different liquids can be discharged simultaneously. Furthermore, it is clear that the dosing heads used according to the present invention can be produced with the aid of arbitrary conventional methods. The dosing head can, for example, be produced from silicon by micromechanical methods. Alternatively, also other known methods, such as micro-injection moulding, hot-process embossing, or methods in the case of which individual layers are adhesively connected or laminated may be used.
The dosage device according to the present invention can be operated either as a dosage device for discharging an amount of liquid predefined by the geometrical volume of the nozzle channel, or as a device having a small but variable volume. In the first case, the driving force acting on the liquid is maintained until the whole amount of liquid contained in the nozzle has been ejected through the nozzle opening. Dosing will stop automatically in this case, since, due to the diminishing pressure gradient, liquid will not be resupplied by the drive unit. In the second case, the driving force applied by the drive unit will be switched off before the liquid has been ejected completely from the nozzle.
It follows that the dosed amount of fluid can be controlled either by the structural shape, i.e. by the volume contained in the nozzle, or by the duration and by the course of the driving force applied by the drive unit. In the first case, the dosed amount of liquid is essentially independent of the physical properties of the liquid, such as viscosity and surface tension. In the second case, the dosed volume will be influenced by these parameters. It is therefore advisable to carry out a calibration in the last-mentioned case so as to achieve a precise dosage, since the dosage of different liquids with different viscosities will take different periods of time. These different periods of time will have to be taken into account if the dosed volume is not the whole nozzle volume, whereas they are of secondary importance in cases in which the whole nozzle volume is discharged, so that the dosing process will stop automatically as soon as the whole amount of liquid corresponding to the predefined volume has been ejected.
While this invention has been described in terms of several preferred embodiments, there are alterations, permutations, and equivalents which fall within the scope of this invention. It should also be noted that there are many alternative ways of implementing the methods and compositions of the present invention. It is therefore intended that the following appended claims be interpreted as including all such alterations, permutations, and equivalents as falling within the true spirit and scope of the present invention.
Number | Date | Country | Kind |
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101 02 152 | Jan 2001 | DE | national |
Filing Document | Filing Date | Country | Kind | 371c Date |
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PCT/EP02/00186 | 1/10/2002 | WO | 00 | 7/7/2003 |
Publishing Document | Publishing Date | Country | Kind |
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WO02/057015 | 7/25/2002 | WO | A |
Number | Name | Date | Kind |
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4537231 | Hasskamp | Aug 1985 | A |
5593290 | Greisch et al. | Jan 1997 | A |
6116297 | Feygin | Sep 2000 | A |
6354471 | Fujii | Mar 2002 | B2 |
Number | Date | Country |
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199 17 029 | Nov 2000 | DE |
Number | Date | Country | |
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20040074557 A1 | Apr 2004 | US |