Method and apparatus for absorbing a medium into a capillary device

An arrangement known from [1] comprises a microtiter plate having a plurality of wells for receiving an analyte.

Such a microtiter plate is used, for example, in a wide variety of medical and biotechnology applications for receiving liquids to be analyzed, for example in the field of DNA analysis.

Usually, a different analyte to be analyzed is introduced into each well and via a pipette, usually a plurality of side by side trained as a so-called Pipettierkamm element, wherein for example in a pipetting comb each a pipette for each well of a row of the microtiter plate is provided with matrix-shaped recesses.

By means of a pipette, in each case an analyte is taken from the corresponding well into which the analyte is filled and into which the pipette is immersed, ie sucked up, on the basis of a vacuum built up in the pipette.

According to the arrangement known from [1], the pipette is in each case coupled via a hose with a pump uniquely associated with the respective pipette, with which the negative pressure is generated, in such a way that the analyte can be sucked in by means of the pump via the corresponding pipette and correspondingly also again, controlled by the pump, can be introduced into the recess.

Such a known microtiter plate has, for example, 96 wells with a size of 8 cm × 12 cm.

However, such a known microtiter plate can basically have any number of, usually up to 384 depressions.

A disadvantage of the arrangement known from [1] can be seen in particular in the fact that, due to the high number of pumps, it is impracticable or even impossible to do so on such a small area of ​​8 cm × 12 cm for each recess of a row, ie for To provide such a high number of pipettes each have their own pump.

Thus, the production of such a pipette comb and thus such an arrangement for receiving liquid analytes is very complicated and expensive.

It should also be noted that in the arrangement known from [1] a peristaltic pump is usually used in each case for aspirating and introducing the analyte out of or into the respective depression.

A considerable disadvantage of this known arrangement is further to be seen in the fact that for the analysis usually a minimum amount of an analyte to be analyzed in the order of 1 ml is required.

Another disadvantage is the fact that the large number of required pumps associated with the arrangement of hoses is very complicated and thus prone to failure.

Furthermore, in [2] a so-called flow-thru-chip ™ is described, by means of which an analysis of the analyte with regard to the existence of biological material in the analyte is known.

The Flow-Thru-Chip ™, an embodiment of an analysis chip, has a plurality of channels through which the analyte is passed through the analysis chip, the surface of the channels each with catcher molecules, generally with molecules containing the correspondingly sought after biological material, whose existence is to be detected in the analyte, preferably covalently bind.

If a DNA strand with a predetermined DNA sequence is to be determined as the biological material in the analyte, DNA capture molecules having a sequence complementary to the DNA sequence to be determined are applied to the surface of such a liquid channel in the flow-thru chip ,

If the DNA material containing the desired DNA sequence is present in the analyte, the DNA strands will bind with the corresponding DNA capture molecules of opposite, ie complementary, sequence.

In general, such an analysis chip is often used for the analysis, ie for the detection of macromolecular biopolymers, which include, for example, proteins or peptides or else DNA strands of a respectively predetermined frequency.

Furthermore, it is known from [3] to produce a membrane of glass or silicon, which has a plurality of pores with a constant diameter of 0.1 .mu.m to 1 .mu.m.

Thus, the invention is based on the problem of receiving a present in a first phase, for example as a liquid or as a gas first medium in a capillary device, wherein the recording takes place in a comparison with the prior art simplified and cost-effective manner.

The problem is solved by the method and the arrangement having the features according to the independent patent claims.

In a method for receiving a first medium which is in a first phase into a capillary device, a negative pressure is generated in the capillary device. With the negative pressure, the first medium is taken up in the capillary device.

The first medium may be in the form of a liquid or a gas.

For example, the first medium may be a liquid to be analyzed, ie an analyte, which is analyzed using the capillary device and an analysis chip coupled to the capillary device, with which the picked-up first medium is brought into contact.

In this case, the analysis chip is, for example, the flow-thru-chip ™ described in [2], wherein biological material may be contained in the liquid channels of the analysis chip and applied to the surface of the liquid channels, wherein the biological material is arranged such that in the first medium contained biological molecules can be bound by catcher molecules.

Thus, for example, as a biological material, DNA capture molecules can be applied in the liquid channels for binding DNA strands contained in the analyte with the DNA sequence complementary to the sequence of the DNA capture molecules.

It is ensured according to the invention that the negative pressure generated in the capillary device is less than a critical pressure, when exercised in the capillary device, a surface tension generated by the first medium or a second medium described later in the capillary device when the first medium completely absorbed by the capillary device would be overcome.

In this way it is prevented that the second medium, which in a second phase, which is different from the first phase, would be taken up in the capillary device after the first medium has been completely absorbed.

Illustratively, this means that in the capillary device, for example by means of a pump control, which controls a negative pressure in the capillary generating pump such, the negative pressure generated in the capillary device is set such that by the negative pressure, the surface tension of the first medium or, if, for example the first medium is gaseous and the second medium is liquid, the second medium in the capillary device is not exceeded.

The critical pressure in the capillary device is given, for example, according to the following procedure:<maths id="math0001" num=""><math display="block"><mrow><mtext>P = 2</mtext><mfrac><mrow><mtext>S</mtext></mrow><mrow><mtext>r</mtext></mrow></mfrac><mtext> .</mtext></mrow></math><img file="EP1161995A2_D0001.tif" /></maths> being with<ul id="ul0001" list-style="bullet" compact="compact"><li>S is the surface tension generated by the first medium in the capillary device when the first medium has been completely taken up by the capillary device,</li><li>r is the radius of a capillary device with a circular base,</li></ul> referred to as.

In the case that the first medium is a liquid, the second medium may be a gas.

Thus, according to this embodiment of the invention, as will be explained in further detail prevents that when the entire liquid has been taken from a container by means of the capillary, no gas, such as air, is absorbed into the capillary, so that the PUMP result this way and associated with it the analysis result when using an analysis chip, is not affected by the second medium.

However, the first medium may also be a gas, in which case the second medium is usually a liquid.

The invention clearly exploits the effect that is automatically ensured by a generated due to the capillary surface tension of the liquid that only the medium to be analyzed is included in the capillary, as by the surface tension after complete absorption of the first medium by the surface tension of the second medium not recorded. If the second medium is in the form of a liquid and the first medium is a gas, then, after complete absorption of the first medium into the capillary device, its absorption is prevented even by the surface tension of the second medium.

This procedure is very simple and the arrangement provided for carrying out the method is thus very inexpensive to produce.

Through the use of an analysis chip for analyzing the medium taken up by the capillary device, an arrangement for the analysis of a medium, for example an analyte for tissue analysis, is made possible overall in a very simple and cost-effective manner.

According to a further embodiment of the invention, the capillary device is a porous plate with a plurality of channels, wherein in each case a channel, the negative pressure is generated.

Each channel has, for example, a circular base area with a radius of 0.1 μm to a few μm, preferably up to approximately 10 μm. In the case of a non-circular cross-section of the channel, the base area is dimensioned in the circular base area of ​​corresponding size.

Embodiments of the invention are illustrated in the figures and will be explained in more detail below.

Show it

<dl id="dl0001"><dt>FIG. 1</dt><dd>a sketch of an arrangement for receiving liquid analytes according to a first embodiment of the invention;</dd><dt>FIG. 2</dt><dd>a section of the arrangement of Figure 1 in cross section in a state in which the entire analyte is in the wells;</dd><dt>FIG. 3</dt><dd>the section of Figure 2 in the state that a portion of the analytes has been sucked by the pipettes into a receiving space;</dd><dt>FIG. 4</dt><dd>a cross-section through a pipette, on the basis of a principle on which the second embodiment of the invention is based is illustrated;</dd><dt>FIG. 5</dt><dd>a cross-section through a pipette, on the basis of a principle on which the second embodiment of the invention is based is illustrated;</dd><dt>FIG. 6</dt><dd>a cross-section through a pipette, on the basis of a principle on which the second embodiment of the invention is based is illustrated;</dd></dl>

First embodiment:

<b>Fig.1</b> shows an arrangement 100 for receiving liquid analytes according to a first embodiment of the invention.

The arrangement 100 has a microtiter plate 101 with a multiplicity of depressions 102 for receiving usually respectively different analytes, ie liquids to be analyzed.

On the microtiter plate 101, a further plate 103 is applied, which is coupled to the microtiter plate 101 by means of screws (not shown). The further plate 103 will be explained in more detail below.

Over the further plate 103, which according to the wells 102 each have pipettes, as shown in Fig. 2, are coupled airtight with an applied on the other plate 103 pump 104.

By means of the pump 104, the pressure within the further plate 103, as described below, adjustable, ie it is freely adjustable in the corresponding space by the pump 104, an overpressure or a negative pressure.

<b>Fig.2</b> shows an enlarged section 105 of the arrangement 100 from <b>Fig.1.</b>

As <b>Fig.2</b> 2, an analyte 201 to be analyzed is usually introduced into the recesses 102.

The pipettes 202 arranged in the further plate 103 are arranged in the further plate 103 in such a manner that when the further plate 103 is mounted on the microtiter plate 102 by means of the screws, not shown, in each case a pipette 202 into a depression 102 assigned thereto and thus into the respective analyte 201 protrudes.

The pipettes 202 are formed on a lower plastic body 203 of the further plate 103.

The lower plastic body 203 is coupled to an upper plastic body 204, for example glued.

According to this exemplary embodiment, it is provided that an intermediate plate 205 is arranged between the lower plastic body 203 and the upper plastic body 204, in which the analysis chip 206, according to this embodiment, the analysis chip described in [2], also called Flow-Thru-Chip ™ is introduced, is such that each one analysis chip 206 is provided in each case for a depression.

Illustratively, this means that in each case one analysis chip 206 is provided for analyzing an analyte 201, which is contained in a depression 102 and according to a method described below via the pipette 202 and the lower plastic body 203 through the analysis chip 206, ie through the liquid channels of the analysis chip 206 is sucked into the upper plastic body 204.

In this way, the analyte 201 is in each case brought into intimate contact with the catcher molecules on the surface of the liquid channels of the analysis chip 206.

On the upper plastic body 204, a diaphragm 207 is provided for a recess 102 in each case.

This means that the upper plastic body 204 each forms a substantially corresponding to the upper surface shape of the recess 102 space which is formed respectively by side walls 208 of the upper plastic body 204.

Clearly, 204 chambers 209 are thus formed in the upper plastic body, which are each bounded by the walls 208, the membrane 207 and the intermediate plate 205 with the integrated analysis chip 206th

The membrane 207 is in each case an elastic membrane, for example made of latex, which can be changed by means of a pressure change in a space 210 located above the upper plastic body 204, which is coupled to the pump 104.

The space 210 may be filled with gas or with a liquid, wherein the membrane for the corresponding gas or the liquid with which the space 210 is filled, is not permeable.

Illustratively, due to a change in pressure in the space 210, the membrane 207 is deformed to produce a pressure change in the respective chambers 209, whereby the analyte 201 is either aspirated or pushed back into the depression via the pipette 202 through the analysis chip 206.

The fluid channels in the Flow-Thru-Chip ™ 206 are coated with biological material, ie DNA capture molecules according to this embodiment, which are bound to the surface of the fluid channels in the analysis chip 206 by the known gold-sulfur coupling.

If the analyte 201 to be analyzed has DNA strands with a sequence that is complementary to the DNA sequence of the DNA capture molecule, these DNA strands covalently bind to the DNA capture molecules in the fluid channel of the analysis chip 206.

Illustratively, the membrane 207 is thus each symbolized by pressure change, as shown in Figure 3, according to the size of the membrane between the two extreme positions, in Figure 3 by the tangents 211, 212 to the respective maximum arched membrane.

Due to the deformation, as described above, the analyte is sucked or discharged.

Further, according to this embodiment, in the lower plastic body 203 for each pipette 202, a bounce plate 213 is respectively provided between the pipette 202 and the intermediate plate 205, by which improved mixing of the analyte 201 is ensured by forming a corresponding flow shape around the baffle plate 213.

It should be noted in accordance with this embodiment that the amount of liquid of the analyte 201 circulated by the membrane 207 should be significantly greater than the volume of a lower chamber 214 defined below by the lower plastic body 203 for each pipette 202 below the analysis chip 206.

After analysis of the analytes, which typically lasts several hours, for example in the context of a hybridization, the arrangement 100 is emptied by means of membrane maximum position in the position 212.

Rinses of the arrangement by means of a rinse solution can be done in a similar manner as the analysis.

Second embodiment:

The second embodiment corresponds essentially to the first embodiment with the difference that no membrane 207 is required.

In order to ensure that, after the entire analyte has been sucked from a respective well, no air or other gas from the depression is sucked into the pipette, the pump 104 is operated such that a surface tension described in the following the lower end of the respective pipette 202 in the analyte is not exceeded.

This principle is in <b>Figure 4</b> illustrated.

<b>Figure 4</b> shows a pipette 401 immersed in a well 402 and thereby in the analyte 403.

A negative pressure formed in the pipette 401 is in <b>Figure 4</b> symbolized by an arrow 404.

The pipette 401 according to this embodiment is configured as a tube with a diameter of about 1 cm and at its lower end 405 with a membrane 406 completed, for example glued, wherein the membrane 406 a plurality of pores 407, but at least one pore 407, with a preferably constant diameter, according to this embodiment has a diameter of 10 microns.

In general, such a pore 407 may, for example, have a diameter of 0.1 μm to 100 μm.

According to this embodiment, a membrane 407 as known from [3] is made of glass or silicon.

According to this embodiment, without limiting the generality, it is assumed that the membrane 407 is made hydrophilic.

The analyte 403 now penetrates into the pores 407 of the membrane 406 and can be sucked into the pipette 401 by a slight negative pressure, according to this exemplary embodiment, for example of 0.03 bar.

If the depression 402 is emptied, ie if the analyte 403 has been completely taken up in the pipette 401, then, as in FIG <b>Figure 5</b> a meniscus 503 is shown at each pore opening 501 between the analyte 403 and the air 502 located only in the depression 402.

In order to deform the forming meniscus 503 such that entry of air 502 into the pore 407 becomes possible, a much greater negative pressure must be generated than the negative pressure required to introduce the analyte 403, generally a liquid, into the capillary , ie in the pipette 401, suck.

This required pressure P can be estimated according to the following instructions:<maths id="math0002" num=""><math display="block"><mrow><mtext>P = 2</mtext><mfrac><mrow><mtext>S</mtext></mrow><mrow><mtext>r</mtext></mrow></mfrac><mtext>.</mtext></mrow></math><img file="EP1161995A2_D0002.tif" /></maths> in which<ul id="ul0002" list-style="bullet" compact="compact"><li>with S the surface tension of the respective    Liquid, ie the analyte 403, and</li><li>with r the radius of the respective pore 407,</li></ul> referred to as.

These quantities are usually known for a given arrangement.

If water is used as the analyte and a pore 407 has a radius of 10 μm, the result for the required pressure P is 0.29 bar.

In order to prevent air entry into the pore 407, it is necessary to ensure a pressure by the pump which is below this estimated pressure.

This control is usually not critical because, as stated above, a vacuum of 0.03 bar is required to aspirate the analyte, which pressure is one order of magnitude lower than the critical pressure at which the surface tension would be overcome and it would be Entry of air into the pore 407 could come.

In other words, this means that the negative pressure P generated in the pipette for this pipette with the above dimensions is in a range of 0.03 &lt;P &lt;0.29 bar.

Thus, air in the pipette can be prevented in a very simple manner.

It is of course also possible for a hydrophobic membrane 407 to pump a predeterminable gas in an analogous manner by means of the arrangement described above and to prevent liquid entry through the respective pore, generally through a capillary.

Clearly, this embodiment makes it possible to detect automatically whether the entire analyte 403 has already been taken up from the respective depression.

It is also automatically ensured that no other medium is included in the analysis device than the material to be analyzed.

<b>Figure 6</b> shows the enlarged section of a lower end of a pore 407 <b>Figure 4</b> at a negative pressure that is within a range that is about to enter air 502 into pore 407.

This is clear due to the strongly curved meniscus 503.

This document cites the following publications:<ul id="ul0003" list-style="none"><li>[1] M. Winter, Robotics and Automation Concepts in Combinatorial Chemistry - Synthesis and Pipetting Robots, Transcript Laborwelt, No. 1, pp. 25-29, 2000;</li><li>[2] A. Steel et al., The Flow-Thru Chip: A Three Dimensional Biochip Platform, Microarray Biochip Technology, edited by M. Schena, pp. 87-117, 2000;</li><li>[3] EP0 296 348 B1</li></ul>