WO2000053318A1

WO2000053318A1 - Test sample card filled in combination with at least a buffer supply

Info

Publication number: WO2000053318A1
Application number: PCT/FR2000/000578
Authority: WO
Inventors: Bruno Colin
Original assignee: Biomerieux S.A.
Priority date: 1999-03-09
Filing date: 2000-03-09
Publication date: 2000-09-14
Also published as: FR2790686B3; AU3295000A; FR2790686A1; EP1156878A1; JP2003517582A; CA2362701A1

Abstract

The invention concerns a test sample card (1) comprising at least: an orifice for introducing a fluid sample in the card (1); an intake compartment (3) for receiving, directly or indirectly, all or part of the sample introduced, and a primary flow path for transporting said sample from the insertion orifice to the intake compartment (3) (direct transfer), or from an intermediate compartment towards said intake compartment (indirect transfer). The invention is characterised in that each intermediate or intake compartment (3) is connected to a buffer supply (5), located in the card (1), which is so arranged and configured prevent the card from being filled by the introduced sample. The invention is particularly applicable to microfluidic devices used in biology.

Description

Analysis card whose filling is associated with at least one buffer volume

DESCRIPTION

The present invention relates to an analysis card comprising at least one opening for the introduction of a fluid sample into the card, an arrival compartment which can receive, directly or indirectly, all or part of the sample introduced, and a primary channel for supplying said sample from the introduction opening to the arrival compartment (direct transfer), or from an intermediate compartment to said arrival compartment (indirect transfer). The invention relates more particularly to means for improving the filling of each compartment.

Document US-A-5, 230,866 proposes a card which corresponds substantially to the technical field of the applicant's invention, but in which the transfer of the sample to the compartment concerned is only carried out indirectly since there is there is always a compartment between the entrance and the compartment concerned. A single element in this invention can be compared to means making it possible to improve the filling, such as a buffer volume according to our invention, it is the large interior chamber, referenced 102. However, this compartment is connected to outside by a capillary channel and an opening. In addition, the point of intersection between the channel, which connects the compartment to the interior wide chamber, and said interior wide chamber does not meet the specifications of the invention of the applicant.

The present invention of the applicant comprises at least one opening for the introduction of the sample. Optionally, there is an opening to the outside, however it is also planned to be able to close this opening by means of a valve.

Other documents, such as for example US-A-5,288,463, propose a simple venting, via a channel of the contents of a compartment.

It is therefore not a real buffer volume within the card, with the advantages of the present invention when it is desired to automatically fill several compartments automatically. The state of the art consists of patents US-A-5, 746,980, US-A-5, 766,553 and US-A-5,843,380, issued to the applicant. These documents disclose an improvement for analysis cards. Thus, the analysis compartments of this card are associated with a bubble trap, which is constituted by a small container connected to the compartment by a channel.

However, this bubble trap is of a volume much less than the volume of the compartment with which it is associated. The trap therefore does not have the role of facilitating the filling of said compartment, but of avoiding the presence of bubbles in the well of the compartment to facilitate optical reading during analysis. In addition, the analytes present in the bubbles which explode in said trap will not be used in the reaction to be carried out in this compartment, which can affect the accuracy of the analysis.

According to the present invention, at least one of the compartments is associated with a container, which is not a bubble trap, since it must not contain liquid. By cons it increases the total volume of said compartment and thus facilitates its filling. Of course it is quite possible to incorporate between the compartment and the container, a bubble trap as defined above.

To this end, the present invention relates to an analysis card comprising at least:

- an opening for the introduction of a fluid sample into the card,

- an inlet compartment which can receive, directly or indirectly, all or part of the sample introduced, and

a primary channel for supplying said sample from the introduction opening to the arrival compartment (direct transfer), or from an intermediate compartment to said arrival compartment (indirect transfer), characterized in that each intermediate or inlet compartment is connected to a buffer volume, located within the card, the arrangement and configuration of which prevent filling with the sample introduced. Preferably, a channel is present between the compartment and the buffer volume.

According to an alternative embodiment, the card is positioned vertically or inclined during its use, and therefore during the transfer of the fluid sample, and the point of intersection between the compartment and the channel is positioned in the upper part, and preferably at the highest level, of this compartment.

According to another additional variant embodiment, this card is positioned vertically during its use, and therefore during the transfer of the fluid sample, and the point of intersection between the buffer volume and the channel is positioned in the upper part, and preferably at the highest level, of this buffer volume.

In the case where the card is of substantially flat shape, which has two opposite faces which are substantially parallel, the compartment is located on one face of the card and the buffer volume is located on the opposite face, said compartment and said buffer volume being connected to each other by a through channel.

In a particular embodiment, the capacity of the compartment and the capacity of the buffer volume are substantially identical.

According to another alternative embodiment, an apparatus, making it possible to break the bubbles that the fluid sample could create, is present at the level of or replaces the channel.

Such a device is already well described in the prior art developed previously, but also in two patent applications filed by the applicant on the same day as the present invention and entitled for the first: "Device and method for positioning a liquid ", and for the second:" Pumping device for transferring at least one fluid into a consumable ".

The content of the description of these patent applications is also considered to be incorporated into the present invention.

Moreover, the buffer volume can be a very important element in the pumping device according to the second patent application described above. So when you press the flexible film to compress the volume of one of the compartments, it is also possible to simultaneously compress the associated buffer volume. It is therefore particularly advantageous to have a compartment and a buffer volume which are situated on two opposite faces of the card, at the same level from one another, so that the pressures on the two containers add up. Of course, it is necessary that each container is covered with a flexible film, which can be the same, provided that said film sandwiches said card.

More specifically, the device, making it possible to break the bubbles, consists of a chamber of section greater than the orifice present between the compartment and the chamber. In the case where the card has two opposite faces, at least one of which is coated with a flexible film, partitioning the card, the buffer volume comprises at least one stiffening tab, in order to prevent the film from coming into contact with the bottom of said buffer volume.

Whatever the embodiment, the card comprises at least: - two arrival compartments which can receive, directly or indirectly, part of the sample introduced, and

- two primary channels for supplying said sample from the introduction opening to the arrival compartments (direct transfer), or at least one intermediate compartment for said arrival compartments (indirect transfer), each primary channel d brought only working with an inlet compartment and the assembly constituting, with the associated buffer volume, a reaction line, said card is characterized by the fact that the arrangement and configuration of each reaction line cause a pressure drop identical to that undergone in each other reaction line in parallel and allowing identical filling of each container.

According to a first alternative embodiment, the buffer volumes are connected to each other.

According to a second alternative embodiment, each buffer volume is constituted by a channel which has at least one opening on the outside of the card. According to a third alternative embodiment, the opening is located at the free end of the channel constituting the buffer volume. According to a fourth alternative embodiment, all of the buffer volumes have a common opening.

According to a fifth alternative embodiment, a valve is present at the opening. According to a particular embodiment, the card comprises at least:

- two inlet compartments which can receive, directly or indirectly, part of the sample introduced, and

- two primary channels for supplying said sample from the introduction opening to the arrival compartments (direct transfer), or at least one intermediate compartment for said arrival compartments (indirect transfer), each primary channel d brought only working with an inlet compartment and the assembly constituting, with the associated buffer volume, a reaction line, said card is characterized by the fact that all the reaction lines comprises a single common buffer volume. According to another alternative embodiment, a buffer volume is associated with each intermediate compartment and / or inlet compartment.

The use of such a card relates to the analysis of one or more different liquid samples in which one seeks to identify one or more analytes according to all the simple or complex analysis processes involving one or more different reagents according to the chemical, physical or biological nature of the analyte (s) sought. The technical principles defined below are not limited to a particular analyte, the only condition being that the analyte is distributed in the sample to be analyzed in suspension or in solution. In particular, the analysis process implemented can be carried out, in homogeneous or heterogeneous or mixed form. One particular, non-limiting mode of such a card relates to the biological analysis of one or more ligands, requiring for their detection and / or their quantification the use of one or more anti-ligands. By ligand is meant any biological species such as, for example, an antigen, an antigen fragment, a peptide, an antibody, an antibody fragment, a hapten, a nucleic acid, a nucleic acid fragment, a hormone, a vitamin. An example of the application of analytical techniques concerns immunoassays, whatever their format, by direct analysis or by competition. Another example of application relates to the detection and / or quantification of nucleic acids comprising all the operations necessary for this detection and / or this quantification from any sample containing the target nucleic acids. Among these various operations, mention may be made of lysis, fluidification, concentration, the steps of enzymatic amplification of nucleic acids, the detection steps incorporating a hybridization step using for example a DNA chip or a labeled probe. Patent application WO-A-97/02357 explains the various steps necessary in the case of analysis of nucleic acids.

The attached figures are given by way of explanatory example and are in no way limiting. They will allow a better understanding of the invention.

FIG. 1 represents a view of one of the faces of an analysis card according to the present invention.

Figure 2 shows a view of the other side of this analysis card.

FIG. 3 represents a sectional view along A-A of FIG. 1 or 2.

Finally, Figure 4 shows a view of the other side of an analysis card according to the present invention, but in a second embodiment.

The present invention relates to a device for facilitating the filling of an analysis card 1, and more particularly of compartments 3, as shown in the figures.

The set of figures represents a particular embodiment in which an analysis card 1 contains five inlet compartments 3. Of course, this number of compartments 3 is absolutely not limiting, and it is possible to have a single compartment 3 as a multitude.

In terms of implementation, the analysis card is obtained by machining a technical plastic material, such as for example impact polystyrene reference R540E from the company GOODFELLOW, compatible with the treated liquids. In an industrial embodiment, the card could be obtained by precision molding, but all other manufacturing methods and in particular those used in semiconductor techniques such as those described in patent application WO-A-97/02357 can be used for the manufacture of analysis cards. The essential objective of this invention is to associate with each container 3 at least one buffer volume 5 which makes it possible to increase the volume available for the aspiration of the liquid sample which must be introduced through the introduction opening 2 or introduction valve. Such a valve 2 is well represented in FIG. 1.

It is noted that from this valve 2, five primary inlet channels 4 leave, allowing the transfer of an introduced biological sample to the inlet compartments 3. In this FIG. 1, the buffer volumes 5 are not shown as will be explained more late. In fact, the card 1 is generally used vertically or possibly inclined. Horizontal use is possible and conceivable, however it is not a preferential mode of use.

Thus, to allow the transfer of the biological liquid from the introduction opening 2 to the inlet compartments 3, it is necessary beforehand to create a depression in the entire pipe network. These techniques are well known to those skilled in the art. For example, it may be a vacuum in the network, as described in a previous patent application filed by the plaintiffs under the number PCT / FR99 / 02082 under priority as of September ^1, 1998.

We note at each compartment 3, the presence of an apparatus 11 which makes it possible to break the bubbles that can be created, when the liquid introduced opens into the container 3. This apparatus 11 essentially consists of a chamber 12 which is separated of the volume of the inlet compartment 3 via an orifice 13, the dimensions of which are smaller than the compartment 3 or the chamber 13. Therefore, any bubble which forms at the level of this orifice 13 comprises an amount of liquid which is not sufficient to accept the increase in section of the chamber 12. The bubbles will therefore explode in this chamber 12.

In the case of vertical use of the card, the liquid present in the bubbles descends to the bottom of compartment 3 after the bubble explodes. There is therefore no loss in line and the filling accuracy of the compartment is improved. This is particularly valid in the case of the use of biological liquids which contain detergents such as Triton XI 00 (registered trademark), Tween 20 (registered trademark), SDS (Sodium Dodecyl Sulfate) or Lauryl Sulfate of Lithium or Sodium and which tend to foam. It is clear from the drawing in FIG. 1 that sharp edges are present at the orifice 13, present between the compartment 3 and the chamber 12 to promote the bursting of the bubbles.

At the level of said chamber 12, we also note the presence of a channel, which is in fact referenced 6 in FIG. 3, of which we only see the point of intersection 7 existing between this channel 6 and the compartment 3, or more exactly between the channel 6 and the apparatus allowing the bubbles 1 to be broken 1. However, one can quite imagine that each compartment 3 does not have an apparatus for breaking the bubbles 11 and that this point of intersection 7 is directly present in the container 3.

Due to the vertical use of the card 1, as has been specified above, the device 11 is located in a position higher than the compartment 3. Similarly, the point of intersection 7 is located in the upper position of the 'device 11. This configuration allows when filling a compartment 3 to avoid filling the buffer volume associated therewith. Finally, in this figure, we note the presence in the lower part of each compartment 3 of an outlet opening, also called outlet valve 19. Valves which can be used in the present invention exist in the state of the art, and also the request Patent filed by the Applicant dated September 8, 1998, under the filing number FR98 / 11383, the content of the description of which is considered to be incorporated into the present application, and the title of which is as follows: "Device allowing reactions , transfer system between devices and method of implementing such a system ”.

The presence of a valve, whether at the level of the introduction 2 or of the outlet 19, makes it possible to control the movement of the liquid sample introduced. In a particular embodiment, where it is necessary to heat a compartment to promote a chemical or biological reaction at temperatures of, for example, between 30 and 120 ° C and advantageously between 40 and 95 ° C, the presence of valves makes it possible to limit the evaporation phenomena which can lead to a modification of the volume in the compartment inducing uncontrolled heating of the reagents or a contamination problem in another compartment. Another advantage of the buffer volume, in this particular embodiment including a heating step, is to absorb the pressure variations due to heating making it unnecessary to have a vent on the analysis card and therefore reducing the risks contamination. For this it is preferable that the buffer volume is thermally insulated or else positioned at a location on the card where the heat transfer due to the heating of the reaction compartment associated with it is minimized.

Of course, one can imagine a channel or several channels allowing the liquid to be brought to the level of the introduction opening 2 and also at least one discharge channel to the level of the outlet opening 19, these associated channels being able to be integrated into said analysis card 1, and allow the liquid to be sent to other areas of the card or other cards, where other treatments and / or reactions may be carried out.

In Figure 2, the other side of the analysis card is shown. The common point between these figures 1 and 2 resides in the presence also of the channel 6 which is symbolized on this face by the point of intersection 8. This point 8 is the point of intersection present between the buffer volume 5 and the channel 6 This channel 6 is well represented in FIG. 3. With regard to FIG. 2, it is noted that the buffer volume is of a fairly large volume, that is to say at least equal to the volume of each compartment d arrival 3, even when the volume of this compartment 3 is associated with the volume of its primary supply channel 4. In this FIG. 2, the shape of the buffer volume is substantially rectangular but other shapes could be chosen without affecting the function of this buffer volume. This buffer volume can vary between 1 and 5000 microliters, advantageously between 5 and 2000 microliters and preferably between 10 and 1000 microliters. The volume of compartment 3 can vary substantially in the same proportions. By way of example, in the case of FIG. 2, a buffer volume, defined by a rectangular parallelepiped of 30 millimeters (mm) by 10 mm and a depth of 1 mm, is shown. The stiffening tabs do not significantly modify the volume. The through channel 6 or the channel inlet 4 can be of section, rectangular, oval or circular. For example, a through channel 6 of circular section with a diameter between 0.1 and 4 mm and advantageously between 0.3 and 2 mm is adapted to the buffer volume described in FIG. 2. In this configuration and always by way of example , the primary inlet channel 4 can vary in the same proportions.

If we now refer to FIG. 3, we note that the analysis card 1 is of flat shape, that is to say that it has two faces, a face 9 where the compartment 3 is located and a face 10 where the buffer volume 5 is located. The link between these two elements 3 and 5 resides in the presence of the channel 6 which is a through channel. The channel is connected to compartment 3, and more particularly to the apparatus for breaking bubbles 11, by means of an intersection point 7 and to the buffer volume 5 by means of an intersection point 8 , already defined previously. The two faces 9 and 10 are covered by a partition or a flexible film 14 which delimits the different volumes.

The nature of the flexible film may vary depending on the nature of the analysis card and of the fluids tested, in particular for compatibility reasons. For example, a TPX (polymethylethylpentene) or BOPP (bi-oriented polypropylene) polymer film makes it possible to carry out biological tests. The fixing of these films can be carried out by gluing (coating of glue such as for example silicone glues on the film) or by welding. An example of adhesive BOPP is provided by the company BioMérieux Inc (St Louis, MO, USA) under the reference 022004-2184.

The dimensions of the buffer volume 5 being quite large, it may be necessary to add stiffening tongues 16, as shown in FIGS. 2 and 3. At the level of compartment 3, it is noted that the introduction of the liquid into said compartment 3 is carried out by means of a drainage means 20 which consists of a slope due to a shape of the bottom of said compartment 3. This particular shape makes it possible to use capillary action when the liquid opens up said compartment and thus to facilitate the liquid spilled into this compartment 3. As can now be seen in FIG. 4, the buffer volume can be made up of different shapes. Compared to FIG. 2, the presence of the intersection points 8 between the buffer volume 15 and the through channel 6 not shown in this figure is again noted. In the case of FIG. 4, each buffer volume 15 consists of a channel which has a suitable length, volume and shape making it possible to control the pressure drop at each compartment 3. Therefore, it is possible to play either on the diameter of the channel or on the number of turns or finally on the length of each of these channels to modulate this pressure drop. The dimensions of the channel constituting the buffer volume 15 are defined by a person skilled in the art. The dimensions given above for channels 6 and 4 can be used, but advantageously, a reduction in size over all or part of the channel to increase the pressure drop effect will be applicable, for example a semicircular section with a diameter of 0.02 to 0 , 6 mm and preferably from 0.2 to 0.4 mm. It is moreover the same state of mind which has given rise to the shape of the primary inlet channels 4 represented in FIG. 1.

A reaction chain is in fact made up of a primary supply channel 4 associated with a compartment 3, possibly with a device making it possible to break the bubbles 11, with a through channel 6 and with a buffer volume 5 or 15, which can have different shapes.

The objective of the present invention is also to allow identical filling of each compartment 3. To do this, it is necessary that the shape of each primary supply channel 4 in combination with the buffer volume 5 or the volume and shape of the buffer volume 15 are calculated to have an identical pressure drop at each compartment 3. Therefore, when the liquid is introduced at the opening 2, the distribution will be completely balanced in each compartment 3. This is particularly interesting in the field of biology where the microfluidics which is used on the charts of analysis can involve, by place, phenomena of retention of liquid. Calculation and proper consideration of these capillarity and pressure drop phenomena make it possible to obtain identical volumes in each compartment 3.

According to embodiments shown in Figure 4, it is possible to connect the buffer volumes together. Thus, the free ends of the buffer volumes 15 meet at the opening 17 of the buffer volume 15 on the outside where the valve 18 is located.

According to an embodiment not shown in the figures, it is also possible that all of the channels 6 all meet in the same buffer volume.

REFERENCES

I. Analysis card 2. Introduction opening or introduction valve

3. Arrival compartment

4. Primary inlet channel

5. Buffer volume

6. Channel or through channel 7. Point of intersection between compartment 3 and channel 6

8. Intersection point between buffer volume 5 and channel 6

9. Face of card 1 where compartment 3 is located

10. Face of the card 1 where the buffer volume 5 is located

I I. Bubble-breaking device 12. Device chamber 11

13. Orifice present between compartment 3 and chamber 12

14. Flexible film

15. Buffer volume

16. Stiffening tab 17. Opening of the buffer volume 15 on the outside

18. Valve associated with the opening 17

19. Outlet opening or outlet valve

20. Means of drainage of a liquid

Claims

1. Analysis card (1) comprising at least: - an opening (2) for the introduction of a fluid sample into the card (1),

- an inlet compartment (3) which can receive, directly or indirectly, all or part of the sample introduced, and

- a primary inlet channel (4) of said sample from the introduction opening (2) to the inlet compartment (3) (direct transfer), or from an intermediate compartment to said inlet compartment (3 ) (indirect transfer), characterized in that each intermediate or arrival compartment (3) is connected to a buffer volume (5 or 15), located within the card (1), the arrangement and configuration of which prevent filling with the sample introduced.

2. Card according to claim 1, characterized in that a channel (6) is present between the compartment ((3) and the buffer volume (5 or 15).

3. Card according to any one of claims 1 or 2, characterized in that it (1) is positioned vertically or inclined during its use, and therefore during the transfer of the fluid sample, and that the point of intersection (7) between the compartment (3) and the channel (6) is positioned in the upper part, and preferably at the highest level, of this compartment (3).

4. Card according to any one of claims 1 to 3, characterized in that it (1) is positioned vertically during its use, and therefore during the transfer of the fluid sample, and that the point of intersection (8) between the buffer volume (5) and the channel (6) is positioned in the upper part, and preferably at the highest level, of this buffer volume (5).

5. Card according to any one of claims 1 to 4, of substantially flat shape, which comprises two opposite faces (9 and 10) which are substantially parallel, characterized in that the compartment (3) is located on one face (9) of the card (1) and that the buffer volume (5) is located on the opposite face (10), said compartment (3) and said volume buffer (5) being connected to each other by a through channel (6).

6. Card according to any one of claims 1 to 5, characterized in that the capacity of the compartment (3) and the capacity of the buffer volume (5) are substantially identical.

7. Card according to any one of claims 1 to 6, characterized in that an apparatus (11), making it possible to break the bubbles that the fluid sample could create, is present at the level of or replaces the channel ( 6).

8. Card according to claim 7, characterized in that the device (11), making it possible to break the bubbles, consists of a chamber (12) of section greater than the orifice (13) present between the compartment (3) and the chamber (12).

9. Card according to any one of claims 1 to 8, at least one of the two opposite faces (9 and / or 10) of which is coated with a flexible film (14), partitioning the card (1), characterized by the fact that the buffer volume (5) comprises at least one stiffening tab (16), in order to prevent the film (14) coming into contact with the bottom of said buffer volume (5).

10. Card according to any one of claims 1 to 9, comprising at least: - two inlet compartments (3) which can receive, directly or indirectly, part of the sample introduced, and - two primary channels d '' (4) of said sample from the introduction opening

(2) to the inlet compartments (3) (direct transfer), or at least one intermediate compartment to said inlet compartments (3) (indirect transfer), each primary inlet channel (4) not operating with an inlet compartment

(3) and the assembly constituting, with the buffer volume (5 or 15) associated therewith, a reaction line, characterized in that the arrangement and configuration of each reaction line cause a pressure drop identical to that undergone in each other reaction line in parallel and allowing identical filling of each compartment (3).

11. Card according to claim 10, characterized in that the buffer volumes (5 and / or 15) are connected to each other.

12. Card, according to any one of claims 10 or 11, characterized in that each buffer volume (15) is constituted by a channel (15) which comprises at least one opening (17) on the outside of the card (1).

13. Card according to claim 12, characterized in that the opening (17) is located at the free end of the channel (15) constituting the buffer volume (15).

14. Card according to claim 11 in combination with any one of claims 12 or 13, characterized in that all of the buffer volumes (5 and / or 15) have a common opening (17).

15. Card according to any one of claims 12 to 14, characterized in that a valve (18) is present at the opening (17).

16. Card according to any one of claims 1 to 9, comprising at least: - two arrival compartments which can receive, directly or indirectly, part of the sample introduced, and

- two primary channels for supplying said sample from the introduction opening to the arrival compartments (direct transfer), or at least one intermediate compartment for said arrival compartments (indirect transfer), each primary channel d 'feed functioning only with an inlet compartment and the assembly constituting, with the associated buffer volume, a reaction line, characterized in that all of the reaction lines have a single common buffer volume.