EP4635613A1

EP4635613A1 - Fluid mixing unit and fluid mixing method

Info

Publication number: EP4635613A1
Application number: EP24171324.7A
Authority: EP
Inventors: Dongwei WU; Jens Kurreck; Shumin Pang
Original assignee: Technische Universitaet Berlin
Current assignee: Technische Universitaet Berlin
Priority date: 2024-04-19
Filing date: 2024-04-19
Publication date: 2025-10-22

Abstract

The disclosure refers to a fluid mixing unit (1), comprising a connector (2) with a first lock section (3) configured to provide a leak-free connection with a first fluid supply container (4) and a second lock section (5) configured to provide a leak-free connection with a second fluid supply container (6), the connector (2) having a receiving room (7) formed between the first lock section (3) and the second lock section (5). The receiving room (7) is respectively sealed against the lock sections (3, 5). The fluid mixing unit (1) further comprises a mixing element (8) extending from the first lock section (3) into the second lock section (5) and through the receiving room (7), the mixing element (8) having an internal fluid passage (9) configured to provide a fluid communication between the first lock section (3) and the second lock section (5), the mixing element (8) being movable in a direction back and forth between the first and the second lock section (3, 5). Further, a method of mixing fluids is provided.

Description

The invention relates to a fluid mixing unit, comprising a connector with a first lock section configured to provide a leak-free connection with a first fluid supply container and a second lock section configured to provide a leak-free connection with a second fluid supply container, the connector having a receiving room formed between the first lock section and the second lock section. The invention further refers to an according method.

Background

Bioprinting is a processing method that utilizes 3D printing technique to spatially organize living cells and biomaterials to create bioengineered constructs. The mixture of cells and biomaterials generally is defined as bioink.
The difference of bioprinting compared to non-biological printing is that 3D bioprinting involves additional complexities, such as the choice of materials, cell types, growth and differentiation factors, and technical challenges related to the sensitivities of living cells and the construction of tissues. The greatest technical challenge in bioink preparation related to the sensitivities of living cells is that the mixing process causes harm to the cells.
A known and widely used method for mixing bioink is the fluid supply container coupler method. Such a fluid supply container couple for example is known from US 2022/ 0 298 471 A1 . A disadvantage of the use of such a fluid supply container coupler is that it is not easy to homogeneously mix viscous bioink. Another disadvantage is that the coupler causes harm to cells during mixing. The conflict of objectives when using the known coupler is therefore that, on the one hand, the cells remain intact with gentle mixing, but only an inadequate mixing result is produced and, on the other hand, the mixing is homogeneous with more vigorous mixing, but the cells are damaged.

Summary

It is therefore the objective technical problem of the invention to further develop a connecting device for coupling fluid supply containers in such a way that, on the one hand, good mixing of the fluids to be mixed is achieved and, at the same time, the cells have a high viability.
The problem is solved by the features of the independent claims. Further advantageous embodiments are defined in the respective dependent claims.
Accordingly, the receiving room is respectively sealed against the lock sections and the fluid mixing unit further comprises a mixing element extending from the first lock section into the second lock section and through the receiving room. The receiving room can be configured to receive the middle portion of the mixing element, such that the middle portion is fixed or movable in a direction between the fluid supply containers. The mixing element has an internal fluid passage configured to provide a fluid communication between the first lock section and the second lock section. Further, the mixing element is movable in a direction back and forth between the first and the second lock section.
The receiving room can be a tubular chamber. The tubular chamber can be a pipe section. The first lock section and the second lock section can be connected to both longitudinal ends of the pipe section. The lock sections can also be formed as pipe sections. The lock sections can have a smaller diameter as the receiving room. The lock sections can be sealed against the receiving room by means of a seal. The lock sections can be screwed onto the tubular chamber. The lock sections can have an internal thread and the tubular chamber can have external threads or vice versa. The lock sections can be formed as Luer connectors for providing a connection with a fluid supply container. Therefore, the mixing unit is suitable for most of the commercially available fluid supply containers, such as syringes.
Accordingly, the connecting sections can have external threads onto which a fluid supply container can be screwed. The mixing element can extend through the receiving room into the first lock section and into to the second lock section. The mixing element can pass through the seal arranged between the receiving room and the lock section.
The fluid mixing unit in particular can be configured for mixing bioink. But it also can be suitable for general mixing liquid/gel/gas in two chambers.
The advantage of the mixing element to be movable is that it enables the mixing of the whole fluid volumes contained in the fluid supply containers. It further makes the mixing of the fluids more efficient. Because the end of the mixing unit protrudes further into the fluid of the opposite fluid supply container when one fluid supply container plunger is pressed, the fluid introduced can flow into the middle of the other fluid instead of just hitting it head-on. This significantly increases the mixing of both fluids.
It may be provided that the range of movement of the mixing element is limited in such a way that the fluid communication between the first lock section and the second lock section is maintained when an end position of the mixing element is reached on both sides. This means that the distance between the fluid openings of the mixing element ends and the mixing chamber is greater than the longitudinal movement space of the mixing chamber within the receiving room.
It can be provided that the mixing element has a stop element in the region of the receiving room which limits the longitudinal range of movement of the mixing element within the receiving room. The stop element can be an element that protrudes radially beyond the contour of the passage opening running through the seal.
It can be provided that the mixing element has a first end portion extending from the receiving room into the first lock section in a sealed manner. It further can be provided that the mixing element has a second end portion extending from the receiving room into the second lock section in a sealed manner.
The mixing unit can also have two fluid supply containers, each of which can be screwed onto one of the ends of the mixing unit. The fluid supply containers can have nozzles. The nozzles can each extend into the lock sections areas. Therefore, the inner diameter of the lock sections can be bigger than the outer diameter of the nozzles. The ends of the mixing element can be configured such that the end can extend into the nozzle of a fluid supply container connected to the mixing unit. Therefore, the inner diameter of the nozzles can be bigger than the outer diameter of the ends of the mixing element. This makes it possible for the ends of the mixing element to be actuated by means of the respective fluid supply container plunger in order to move the mixing element in one direction or the other.
The mixing element can further have a middle portion located between the end portions and within the receiving room. The middle portion can have at least sectionally an outline projecting beyond the outlines of the end portions and thus functions as the stop element. The middle portion can have a longitudinal length which is less than a longitudinal length of the receiving room, so that the mixing element can be longitudinally movable inside the receiving room between two end positions. The middle portion can have a mixing chamber which is fluidically fed by the end portions. At least one fluid guiding element can be arranged in the mixing chamber, which forces fluid flowing into the mixing chamber at least sectionally into a flow direction which deviates from the inflow direction. The fluid guiding element can be formed helically. The opposite end faces of the mixing chamber can each serve as stop elements, which in their respective end positions abut against the chamber wall on the end face.
The mixing unit can be configured such that when the middle portion of the mixing element is in a position between the end positions, both end portions of the mixing element protrude beyond the respective lock section. The mixing unit further can be configured such that when the middle portion of the mixing element is in one of the end positions, at least the end portion of the mixing element protrudes beyond the lock section to which the middle portion is approximated.
The mixing element for example can have a length of about 6 cm, suitable for 3 mL fluid supply containers. However, the length of the mixing element can be adapted to fluid supply containers having a different length. The mixing unit can efficiently be used independently from the diameter of the fluid supply containers.
Each end portion can have at least one fluid opening through which a fluid contained in a fluid supply container connected to the mixing unit can flow into the mixing element. The at least one opening can be arranged laterally at the respective end portion. Each end portion also can have two fluid openings. The openings can be arranged laterally opposite each other.
The mixing unit can be compatible with most of the bioinks, as long as the bioinks can go through the fluid openings on the mixing element. Therefore, the viscosity can be from low to very high or the opening size can be adjusted or adapted to the desired viscosity. Therefore, the mixing unit is not only suitable for preparing bioinks of extrusion-based printing (generally higher viscosity), but is also able for bioink mixing for light curing printing (generally lower viscosity).
The invention further concerns a method of mixing fluids, in particular by means of a bioink mixing unit according to any one of the preceding claims, comprising the steps of:

connecting a first fluid supply container containing a first fluid to be mixed to a first lock section of a connector, and connecting a second fluid supply container containing a second fluid to be mixed to a second lock section of the connector so that the fluid supply containers are in fluid communication with each other by means of a mixing element received within the connector;
alternately actuating the fluid supply container plunger of the first fluid supply container so that fluid moves through the mixing element toward the second fluid supply container and actuating the fluid supply container plunger of the second fluid supply container so that fluid moves through the mixing element toward the first fluid supply container by means of the mixing element,
wherein the mixing element moves at least partially in the respective actuating direction in the course of actuating the fluid supply container plungers.

It can be provided that actuating the fluid supply container plunger means moving the fluid supply container plunger to a longitudinal end position in the fluid supply container in which the plunger is maximally approximated to the connector. It is possible that no more fluid is contained in the fluid supply container when the plunger is in its end position.
The connector can have a receiving room in which a middle portion of the mixing element is received, the middle portion can be longitudinally displaceable between a first end position and a second end position within the receiving room.
The middle portion of the mixing element can be in the second end position when the fluid supply container plunger of the first fluid supply container is in its end position. The middle portion further can be in the first end position when the fluid supply container plunger of the second fluid supply container is in its end position. When the middle portion of the mixing element is located in its end position, the stop element can be in contact with the spacer. Alternatively, there still can be a gap between the stop element and the spacer, for example 1 mm, when the middle portion of the mixing element is located in its end position.

Description of further embodiments

Additional details of the invention are explained with reference to the figures below, in which:

Fig. 1: shows a perspective view of the fluid mixing unit according to a first embodiment of the invention;
Fig. 2: shows an exploded view of the fluid mixing unit according to the first embodiment of the invention;
Fig. 3: shows a cross-sectional view of the fluid mixing unit according to the first embodiment of the invention in a first end position of the mixing element;
Fig. 4: shows a cross-sectional view of the fluid mixing unit according to the first embodiment of the invention in a second end position of the mixing element;
Fig. 5: shows a perspective view of the fluid mixing unit according to a second embodiment of the invention;
Fig. 6: shows a detailed view of a fluid guiding element;
Fig. 7: shows a second embodiment of a mixing element;
Fig. 8: shows an application example of the mixing unit, where two syringes are fluidically coupled by means of a fluid mixing unit according to the first embodiment of the invention;
Fig. 9: shows a comparison of the mixing results of a normal Luer coupler compared to the fluid mixing unit according to the first embodiment of the invention;
Fig. 10: shows a comparison of the printing effects during an earlier and a later printing period of a normal Luer coupler compared to the fluid mixing unit according to the invention;
Fig. 11: shows a comparison of the cell viability after one day and after seven days of a normal Luer coupler compared to the fluid mixing unit according to the invention;
Fig. 12: is a diagram showing the cell viability after one day and after seven days of a normal Luer coupler compared to the fluid mixing unit according to the invention;
Fig. 13: shows the relative cell viability in different wells after mixing with normal coupler and mixing unit.

The embodiment of the fluid mixing unit 1 shown in Fig. 1 comprises a connector 2 with a first lock section 3 configured to provide a leak-free connection with a first fluid supply container (not shown) and a second lock section 5 configured to provide a leak-free connection with a second fluid supply container (not shown). The connector 2 has a receiving room 7 formed between the first lock section 3 and the second lock section 5, whereby the receiving room 7 is respectively sealed against the lock sections 3, 5, preventing fluid from entering the receiving room 7. A mixing element 8 is accommodated in the connector 2 and extends through it from the first lock section 3 through the receiving room 7 into the second lock section 5. The mixing element 8 has an internal fluid passage 9 configured to provide a fluid communication between the first lock section 3 and the second lock section 5. The mixing element 8 is movable within in a longitudinal direction back and forth between the first and the second lock section 3, 5 and within the limits defined by the receiving room 7. The mixing element 8 further has a first end portion 11 extending from the receiving room 7 in and/or through the first lock section 3. A second portion 12 of the mixing element 8 extends opposite the first end portion 11 from the receiving room 7 in and/or through the second lock section 5.
To create the fluid passage between the first and second fluid supply containers 4, 6, the end portions 11, 12 each have two lateral fluid openings offset longitudinally in relation to each other, through which the fluids 23, 24 can flow into the mixing element 8. For the mixing process, a first fluid supply container 4 can be connected to the first lock section 3 and a second fluid supply container 6 to the second lock section 5. The fluid supply containers 4, 6 also each have a complementary Luer connector, i.e. female in the example shown, into which a nozzle of the fluid supply container extends in the direction of the mixing unit 8. The mixing element end portions 11 and 12 are again configured to project into the nozzles of the fluid supply containers 4, 6 when extended. For mixing, the fluid supply container plungers 17 are actuated alternately in order to force the fluids 23, 24 in the direction of the mixing unit 1. At a certain point, the plungers 17 collide with the respective end portion 11, 12 of the mixing element 8, so that this moves with the fluid supply container plunger 17 as it continues to move in the direction of the opposite fluid supply container. As a result, the mixing element 8 including the middle portion 13 containing the mixing chamber 14 is longitudinally displaced within the receiving room 7. Similarly, the opposite end portion of the mixing element 8 is moved into the opposite fluid supply container.
Fig. 2 shows an exploded view of a mixing unit 1. This has a connector 2, which consists of a pipe section 26 and screw caps 27 screwed to it on opposite sides, which form the receiving room or chamber 7 when assembled. The outward-facing ends of the screw caps 27 represent the lock sections 3 and 5, which are designed as Luer connectors in the embodiment shown. The mixing element 8 essentially has two rod-shaped end portions 11, 12 and a middle portion 13 arranged between them, which represents the mixing chamber 14. The mixing element 8 extends essentially longitudinally within the connector 2, with the end sections 11, 12 extending away from the mixing chamber 14. In order to seal the lock sections 3, 5 fluid-tight from the receiving room 7, seals 18 are provided between the screw caps 27 and the pipe section 26, through which the end portions 11, 12 extend and are movable in the longitudinal direction through the seal 17. Furthermore, spacers 19 are provided on both sides, which serve as a stop for the end faces of the mixing chamber 14, which act as stop elements 10, when these are in the respective end positions.
Figures 3 and 4 each show cross-sectional views of the mixing unit 1, with the mixing element 8 in the left end position in Fig. 3 and in the right end position in Fig. 4. In the end positions, the end faces or the stop elements 10 of the mixing chamber 14 are in contact with the stops 19 longitudinally delimiting the receiving room 7. The laterally arranged fluid openings 16 at both end portions 11, 12 of the mixing element 8 are clearly visible in the views shown. The fluid guiding element 15 inside the mixing chamber 14 can also be seen. The arrangement of the holes 16 and the design of the fluid guiding element 15 force the fluids 23, 24, which basically move longitudinally, in a transverse direction, which improves mixing.
The second embodiment of the fluid mixing unit 1 shown in Fig. 5 differentiates from the first embodiment shown in Fig. 1 in that the receiving room 3 which in particular accommodates the middle portion 8 of the mixing element 8 is not fully enclosed but configured in an open design, in which the receiving room is not a closed chamber, but in which the lock sections 3, 5 of the mixing unit 1 are spaced apart by means of a plurality of webs 28.
Fig. 6 shows a detailed view of the fluid guiding element 15, which is helical in the embodiment shown. This design forces the fluids onto a path of movement that deviates from the main longitudinal direction of movement.
Fig. 7 shows a second embodiment of the mixing element 8, where the middle portion 13 is spiral-shaped. The first end portion 11 and the second end portion 12 keep the same, but the middle portion 13 is replaced with a compressible spinal structure. In use, this compressible middle portion 13 is accommodated in the receiving room 7. However, the middle portion 13 can be fixed or movable inside the receiving room. For mixing, the plungers 17 of the fluid supply containers 4, 6 force the fluids to flow in the direction of the mixing unit 1. At a certain point, the plungers 17 collide with the respective end portion 11, 12 of the mixing element 8, so that this end portion 11, 12 moves with the plunger 17 as it continues to move in the direction of the opposite fluid supply container 4, 6. For the case that the middle portion 8 is fixed within the receiving room 3, the end portion 11, 12 is pushed by the plunger 17, and the middle portion is compressed to be shorter. For the case that the middle portion 8 is movable, the end portion 11, 12 is pushed by the plunger 17, the middle portion 8 is firstly compressed, and then moved when the pushing force is larger than the resistance. Similarly, the opposite end portion 11, 12 of the mixing element 8 is moved into the opposite fluid supply container 6. When the plunger 17 is pushed on another side, the fluid flows through the mixing element 8 in the opposite direction, and the compressed middle portion 8 is released.
Fig. 8 shows the application of the mixing unit 1. The nozzle 29 of a first syringe 4 is coupled to the first lock section 3 via a Luer connection and the nozzle 29 of a second syringe 6 is coupled to the second lock section 5 of the mixing unit via another Luer connection. The first syringe 4 contains a first fluid 23 and the second syringe 6 contains a second fluid 24, which are to be mixed. By alternately pressing the syringe plungers 17, the fluids are each displaced in the direction of the other syringe respectively, passing through the mixing element 8 and are in particular mixed inside the mixing chamber 14 by means of the fluid guiding element 15. The mixing element 8 is displaced along the mixing unit 1 within its range of movement. Depending on the selected dimensions of the mixing element 8 and/or the receiving room 7, the movement of the mixing element 8 can be stopped either by the end face of the mixing element 8 coming to rest against the opposite stop 19, or by the injection plunger 17 having reached its end position, so that further displacement of the end section 11, 12 is not possible.
Fig. 9 shows the mixing result of two fluids 23, 24 using a conventional Luer connection 25 on the one hand (above) and the mixing result using the fluid mixing unit 1 according to the invention on the other (below). It can be clearly seen that the mixing of the two fluids 23, 24 is significantly more homogeneous when using the mixing unit 1, whereas when using a normal Luer connector as shown above, the two fluids 23, 24 are still more or less in separate phases.
Fig. 10 shows a comparison of the printing effect using the example of a normal Luer 25 on the one hand (left) and the mixing unit 1 on the other (right). The results in an earlier printing period and the results in a later printing period are compared. It can be seen that the results of both connectors are still relatively similar in the earlier stage and predominantly correspond to the desired pattern, although the results of the mixing unit already show fewer defects here. However, in the later stage, a very clear difference can be seen between the results of the mixing unit 1 and those of the normal connector. While the results of the mixing unit still predominantly correspond to the desired pattern, the results of the normal Luer connector 25 show clear defects, no uniform pattern can be recognized any more, but the results all show different forms of clumped material. A similar result as in Fig. 10 is shown in Figures 11 and 12. Fig. 11 shows a live/dead assay of a normal Luer coupler 25 compared to the inventive mixing unit 1 after one day and after seven days. Fig. 12 shows an XTT assay diagram showing the absorbance (A450nm-A620nm) over time from a normal Luer coupler 25 and from the mixing unit 1. The printing results of Figs. 8-10 were generated using the following specifications: bioink with 3% gelatin, 2% alginate, 50 mM CaSO4, 10^7 /mL HepaRG cells, incubated for 8 min at RT, then bioprinted with a mechanical printhead on regenHU 3D discovery bioprinter, 25G needle.
Figure 13 shows the cell viability in different wells from beginning to the end in the cartridge after mixing and bioprinting. The relative cell viability was mainly influenced by cell numbers here. It can be seen that there were fewer cells in the beginning area of the cartridge than the ending area after mixing with a normal coupler 25. However, the cells were distributed much more homogeneously after mixing with the mixing unit 1, showing fairly constant level of viability. Therefore, the inventive mixing unit 1 has the ability to improve the homogeneity during bioink preparation in both transverse and longitudinal directions.
The features of the invention disclosed in the preceding description, in the drawings and in the claims may be essential for the implementation of the invention, both taken in isolation or in various combinations thereof.

Reference numerals

1: Fluid mixing unit
2: Fluid supply containerconnector
3: First lock section
4: First fluid supply container
5: Second lock section
6: Second fluid supply container
7: Receiving room
8: Mixing element
9: Fluid passage
10: Stop element
11: First end portion
12: Second end portion
13: Middle portion
14: Mixing chamber
15: Fluid guiding element
16: Fluid opening
17: Plunger
18: Seal
19: Spacer
20: Luer connector
21: External thread
22: Internal thread
23: First fluid
24: Second fluid
25: Normal Luer connector
26: Pipe section
27: Screw cap
28: Bar
29: Nozzle

Claims

Fluid mixing unit (1), comprising:
- a connector (2) with
- a first lock section (3) configured to provide a leak-free connection with a first fluid supply container (4) and a second lock section (5) configured to provide a leak-free connection with a second fluid supply container (6), the connector (2) having a receiving room (7) formed between the first lock section (3) and the second lock section (5), characterized in that the receiving room (7) is respectively sealed against the lock sections (3, 5); and with

- a mixing element (8) extending from the first lock section (3) into the second lock section (5) and through the receiving room (7), the mixing element (8) having an internal fluid passage (9) configured to provide a fluid communication between the first lock section (3) and the second lock section (5), the mixing element (8) being movable in a direction back and forth between the first and the second lock section (3, 5).
Fluid mixing unit (1) according to claim 1, wherein the range of movement of the mixing element (8) is limited in such a way that the fluid communication between the first lock section (3) and the second lock section (5) is maintained when an end position of the mixing element (8) is reached on both sides.
Fluid mixing unit (1) according to claim 1 or claim 2, wherein the mixing element (8) has a stop element (10) in the region of the receiving room (7) which limits the longitudinal range of movement of the mixing element (8) within the receiving room.
Fluid mixing unit (1) according to claim 3, wherein the mixing element (8) has a first end portion (11) extending from the receiving room (7) into the first lock section (3) in a sealed manner, and wherein the mixing element (8) has a second end (12) portion extending from the receiving room (7) into the second lock section (5) in a sealed manner.
Fluid mixing unit (1) according to claim 4, wherein the mixing element (8) further has a middle portion (13) located between the end portions (11, 12) and within the receiving room (7), the middle portion (13) having at least sectionally an outline projecting beyond the outlines of the end portions and thus functions as the stop element (10).
Fluid mixing unit (1) according to claim 4 or claim 5, wherein the middle portion (13) has a longitudinal length which is less than a longitudinal length of the receiving room (7), so that the mixing element (8) is longitudinally movable inside the receiving room (7) between two end positions.
Fluid mixing unit (1) according to one of the claims 4 to 6, wherein the middle portion (13) has a mixing chamber (14) which is fluidically fed by the end portions (11, 12) and in which at least one fluid guiding element (15) is arranged, which forces fluid flowing into the mixing chamber (14) at least sectionally into a flow direction which deviates from the inflow direction.
Fluid mixing unit (1) according to claim 7, the fluid guiding element (15) being formed helically.
Fluid mixing unit (1) according to claim 7 or claim 8, the mixing unit (1) being configured such that
- when the middle portion (13) of the mixing element (8) is in a position between the end positions, both end portions (11, 12) of the mixing element (8) protrude beyond the respective lock section (3, 5); and

- that when the middle portion (13) of the mixing element (8) is in one of the end positions, at least the end portion (11, 12) of the mixing element (8) protrudes beyond the lock section (3, 5) to which the middle portion (13) is approximated.
Fluid mixing unit (1) according to one of the claims 4 to 9, wherein each end portion (11, 12) has at least one fluid opening (16) through which a fluid contained in a fluid supply container (4, 6) connected to the mixing unit (1) can flow into the mixing element (8).
Fluid mixing unit (1) according to claim 10, wherein the at least one opening (16) is arranged laterally at the respective end portion (11, 12).
Method of mixing fluids, in particular by means of a fluid mixing unit according to any one of the preceding claims, comprising the steps of:
- connecting a first fluid supply container (4) containing a first fluid to be mixed to a first lock section (3) of a connector (2), and connecting a second fluid supply container (6) containing a second fluid to be mixed to a second lock section (5) of the connector (2) so that the fluid supply containers (4, 6) are in fluid communication with each other by means of a mixing element (8) received within the connector (2);

- alternately actuating a plunger (17) of the first fluid supply container (4) so that fluid moves through the mixing element toward the second fluid supply container (6) and actuating a plunger (17) of the second fluid supply container (6) so that fluid moves through the mixing element (8) toward the first fluid supply container (4) by means of the mixing element (8),
wherein the mixing element (8) moves at least partially in the respective actuating direction in the course of actuating the fluid supply container plungers (17).
Method of claim 12, wherein actuating the plunger (17) means moving the plunger (17) to a longitudinal end position in the fluid supply container (4, 6) in which the plunger (17) is maximally approximated to the connector (2).
Method of claim 12 or 13, wherein the connector (2) has a receiving room (7) in which a middle portion (13) of the mixing element (8) is received, the middle portion (13) being longitudinally displaceable between a first end position and a second end position within the receiving room (7).
Method according to claim 14, wherein the middle portion (13) of the mixing element (8) is in the second end position when the plunger (17) of the first fluid supply container (4) is in its end position, and wherein the middle portion (13) of the mixing element (8) is in the first end position when the plunger (17) of the second fluid supply container (6) is in its end position.