WO2012131140A1

WO2012131140A1 - Method and device for applying a liquid in at least one well of a plate

Info

Publication number: WO2012131140A1
Application number: PCT/ES2012/070216
Authority: WO
Inventors: Antonio Alonso Ortiz; Abelardo CABALLERO GONZÁLEZ; Jose María MIRANDA SAYAGO
Original assignee: Servicio Andaluz De Salud; Universidad De Málaga
Priority date: 2011-03-31
Filing date: 2012-03-30
Publication date: 2012-10-04
Also published as: ES2389338A1; ES2389338B1

Abstract

The invention relates to a device that makes use of the viscosity and surface tension properties of liquids in order to fill at least one well in a well plate, while the plate is kept in the inverted position. The invention also relates to a method for filling at lest one well of a plate with a liquid, using the aforementioned device.

Description

PROCEDURE AND APPLIANCE FOR APPLYING A LIQUID IN AT LEAST ONE POOL OF A PLATE

OBJECT OF THE INVENTION

The main object of the invention is a novel apparatus that takes advantage of the properties of the viscosity and surface tension of the liquids to allow the filling of at least one well of a well plate while maintaining it in an inverted position.

Another object of the invention is a method for filling at least one well of a plate with liquid using the above apparatus.

BACKGROUND OF THE INVENTION

For carrying out cultures and tests of different types, plates with a plurality of wells are known (there are, for example, 96-well plates). Normally, the first step to perform the culture or test in question is to fill each well of the plate with a certain liquid. This process is extremely tedious, since it is usually done manually by depositing a drop per well with the help of a dropper. Another drawback of this method is that a lot of liquid is spilled, soiling the surface of the table on which it is carried out, and also in no way equal volumes are obtained in all the wells: while some spill the others stay halfway fill in.

DESCRIPTION OF THE INVENTION

The present invention solves the above problems thanks to a novel apparatus where the application of the liquid is carried out with the plate in an inverted position, taking advantage of the predominant effect of the surface tension over small distances so that the liquid adheres to the inner surface of the well or wells. A great advantage of this device is that it they manage to fill all the wells evenly with liquid in a much faster and cleaner way. Alternatively, if only one well of each plate is filled, said single well of several consecutive plates is filled with the same amount of liquid.

Therefore, the present invention is generally directed to a method for applying a liquid to at least one well of a plate, the method comprising the steps of: placing the plate in an inverted position; and apply the liquid to each well from under said plate.

Furthermore, since the viscosity and surface tension of any liquid are temperature dependent, the apparatus of the invention preferably further comprises means for modifying the temperature of the liquid prior to its application in the wells. In this way, it is possible to control the volume of liquid that is adhered to the well or wells: at a higher temperature, less liquid is adhered to the wells.

Note that in this document, the term "liquid" is intended to refer not only to liquids themselves, but in general any substance capable of flowing sufficiently to be driven with a pump and passing through the needle ducts.

A first aspect of the invention is directed to an apparatus for applying a liquid to at least one well of a plate, where the apparatus comprises means for applying the liquid while the plate is in an inverted position. Preferably, said means basically comprise: a reservoir that stores the liquid, a pump that drives it, and a needle array that applies the liquid to the wells. Each of these elements is described in more detail below:

Deposit It is a reservoir that stores the liquid that you want to apply to the plate. Since not all of the liquid pumped through the needles is adhered to the wells and therefore falls back into the reservoir, the upper surface of said reservoir comprises at least one return hole in its upper surface. This hole could have any shape, provided its size is sufficient to allow the return of all the liquid not adhered to the wells, thus preventing it from dripping down the sides of the apparatus. As for its location and number, in principle it could also be anyone. For example, a reservoir with an inclined upper surface or shaped so as to force the liquid to be directed towards a single return orifice could be conceived. Another solution, corresponding to a preferred embodiment of the invention, is to configure the hole or holes so that they substantially surround the needle array, so that the liquid has no other way than to return to the reservoir.

Preferably, the apparatus of the invention further comprises a support flange for the well plate, thus facilitating proper placement of the plate on the reservoir (that is, so that each needle is inserted into the corresponding well).

Needle Matrix

This is the element that carries out the effective application of the liquid in the wells. In this document, the term "needles" is intended to refer not only to hypodermic needles themselves, but in general to any thin conduit, made of any material, which allows the application of liquid under the conditions described in this application. In addition, it is understood that the most basic "matrix" could be constituted by a single needle (in case the apparatus was configured to fill a single well of a plate).

The needles are arranged vertically with the tip at their end upper, forming a matrix on the upper surface of the tank and the position of the needles corresponding to the position of the wells to be filled with liquid when the plate is correctly positioned. The lower ends of the needles are connected by their base to the reservoir, thus allowing the liquid in the reservoir to rise through the needle ducts until they exit through the tip thereof.

According to a preferred embodiment of the invention, the apparatus of the invention further comprises a protection piece parallel to the upper surface of the reservoir and provided with an array of holes configured for the passage of the tips of the needle array. Thus, the base of the needles is fixed to the upper surface of the tank, but the protection piece serves as protection to them, leaving only the tip visible. In addition, these holes have a clearance around the tips of the needles to allow the passage of the return liquid into the reservoir.

Bomb

It is a suitable pump to propel the liquid from the tank, causing it to rise through the inner ducts of the needles until it comes out through the tips of the needles. Thus, when a well plate is placed in an inverted position on top of the needle matrix, the tip of the needles being in contact with, or very close to, the bottom of each well, the pumped liquid is adhered to the wells by the effect of surface tension. As it is pumped, the liquid accumulates in the wells until the gravitational forces exceed the forces due to surface tension, at which point the excess liquid begins to detach from the well and drip into the tank through of the hole or return holes described above.

There are different possible configurations for the pumping system in general, and the pump itself in particular. The bomb, for example, could be submerged inside the tank. Another option would be to place the pump next to the tank, thus facilitating maintenance tasks. In any case, the pump is preferably connected by means of a feed line to an inner sub-tank comprised within the tank, this being its inner b-tank connected in turn directly to the needles. Thus, when the pump is operated, the liquid rises through the supply line causing an increase in the pressure in the sub-tank, which in turn causes the liquid to rise through the needle lines until it exits the tip, being adhered to the wells as explained above.

There are different ways to control the flow of liquid pumped to the needles. For example, the pump itself could be adjustable. Another possibility, according to a preferred embodiment of the invention, is that the feed duct has an adjustable lateral escape route. Thus, when it is desired to reduce the pumped flow, it is sufficient to open the exhaust path, thus leaving part of the pumped liquid inside the tank before reaching the sub-tank. If it is desired to increase the flow, it would be sufficient to close the escape route to the desired extent to reduce the volume of liquid escaping to the tank.

In addition, as mentioned above, the variation of viscosity and surface tension with temperature can be used to modify the volume of liquid that adheres to the well. In fact, the volume of liquid that adheres to the well depends on the moment in which the balance between surface tension forces and gravitational forces is reached: as the volume of liquid adhered to the well increases, the forces also increase gravitational until reaching a time when excess liquid falls, returning to the tank through the return hole. Taking into account that viscosity and surface tension decrease with increasing temperature, heating the liquid has the effect of decreasing the volume of liquid that adheres to the well. Accordingly, in a preferred embodiment of the invention the apparatus comprises means for heating the liquid contained in the reservoir, preferably a resistor. Moreover, according to another preferred embodiment, the apparatus comprises a thermostat adjustable from outside the tank to control the heating temperature.

This procedure and apparatus could be used in principle with any liquid, although the higher the surface tension of the liquid, the greater the volume trapped in the wells. In addition, it is evident that it would be applicable to plates with any number of wells. Moreover, it would be possible to configure the arrangement of the needles and different support flanges to achieve a versatile apparatus capable of filling plates of different sizes.

Finally, preferably the pump is in continuous operation, so that the technician only has to successively place plates on the apparatus, wait for the liquid to begin to fall into the tank (which is a sign that the wells are full) and Go to the next plate.

A second aspect of the invention is directed to the operating procedure of this novel apparatus, which essentially comprises the steps of:

placing a plate in an inverted position on the needles of the apparatus such that each of the needles is inserted into one of the wells; Y

Actuate the pump to cause the oil to rise through the feed line, reach the sub-tank and ascend through the needles until they exit through the tip of the same to be adhered to the wells of the plate. Preferably, this method comprises the additional step of activating the resistance to control the amount of liquid that is adhered to the wells of the plate. BRIEF DESCRIPTION OF THE FIGURES

Fig. 1 shows a sectional view of an example of the apparatus of the present invention filled with oil.

Fig. 2 shows a sectional view of a conventional well plate.

Fig. 3 shows a sectional view of an example of the apparatus of the invention during the application of oil to a well plate.

Fig. 4 shows a sectional view of the well plate when the final volume of oil in each well has been reached, returning the excess oil to the reservoir.

PREFERRED EMBODIMENT OF THE INVENTION

An example of the apparatus (1) of the invention is described below with reference to the attached figures. In particular, in Fig. 1 the parts comprising the apparatus (1) of the invention can be seen in the rest position: a tank (2) whose upper part is provided with return holes (3); an array of needles (4) that communicate with a sub-tank (10), which is in turn connected by a conduit (9) for communication with the pump (5). It can be seen how the communication conduit (9) has an escape route (1 1), which is shown open in Fig. 1, which will allow to regulate the flow of oil that is pumped into the needles (4).

The upper part of the apparatus (1) also has a protection piece (6) that covers the upper part of the tank (2) and the base of the needles (4), and which is provided with an array of holes (7) of passage of the tips of the needles (4). It is observed how it exists between the holes (7) and the needles (4) a distance that allows the oil that falls from the plate to re-enter the tank (2). In addition, support flanges (8) allow the well plate (100) to be properly placed (101) to be filled in: it is ensured that the position of the wells (101) corresponds to that of the needles (4), and that the tip of the needles (4) is in contact, or almost, with the bottom of the wells (101).

Also shown in the figures is a heating means (12), in this example a resistor, which will serve to heat the oil and thus modify the amount of it that is adhered to the plate

(100) The resistor is connected to a thermostat (13) that allows to regulate the temperature of the oil from the outside of the device simply by turning a wheel. Fig. 2, on the other hand, shows a conventional well plate (100) (101).

Thus, as shown in Fig. 3, when the pump (5) is running, the oil rises through the feed line (9), reaches the sub-tank (10) and ascends through the needles (4) until you exit the tip. When a plate (100) is placed upside down on the needles (4), each of the needles (4) is inserted into one of the wells

(101) and touching, or almost, the bottom. The oil that rises through each needle (4) thus begins to adhere to the wells (101), gradually increasing the oil volume of each well (101). At a given time, oil stops accumulating, either because the well (101) is completely full (as shown in Fig. 4), or earlier because the balance between the surface tension forces and the gravitational forces. At that time, the excess oil begins to slide down the needles (4) down or drip from the wells (101), is introduced through the holes (7) of the protective piece (6) and ends up returning to the tank through the return holes (3). If it is desired to increase the oil flow, it is enough to gradually close the exhaust path (1 1) until the desired flow is achieved. On the other hand, if it is desired to decrease the amount of oil adhered to each well (101) at the end of the process, simply turn on the thermostat (13) to activate the resistance (12). When the oil temperature rises, the surface tension drops and therefore the amount of oil adhered to the wells (101) at the end of the process will also be lower. Thus, playing with the oil temperature, it is possible to regulate the amount administered to each well (101) according to each particular application.

Claims

one . Apparatus (1) for applying a liquid in at least one well (101) of a plate (100) comprising means for applying the liquid in the wells (1 01) with the plate (1 00) being in an inverted position, where said means include:

a liquid reservoir (2) provided with at least one return hole (3) in its upper surface;

a matrix of needles (4) d vertically disposed on the upper surface of the reservoir and whose position is corresponding to the wells (101) that it is desired to fill with liquid, being internal ducts of the needles connected to the reservoir (2) of liquid ; Y

a pump (5) configured to propel the liquid from the reservoir (2) causing its rise through the inner ducts of the needles (4) to exit through the tips of said needles, such that placing a plate (100 ) inverted on top of the needle matrix (4) it is achieved that a portion of the liquid is adhered to the wells (101) by the effect of surface tension, returning the rest of the liquid to the inside of the tank (2) through the minus a return hole (3).

2. Apparatus (1) according to claim 1, wherein the at least one return hole (3) is arranged so that it substantially surrounds the needle array (4).

3. Apparatus (1) according to any of claims 1-2, wherein the reservoir (2) has a flat top surface.

4. Apparatus (1) according to any of claims 1 -3, further comprising a protection piece (6) parallel to the upper surface of the reservoir (2) and provided with an array of holes (7) configured for the passage of the tips of the needle matrix (4).

5. Apparatus (1) according to any of claims 1-4, further comprising a support flange (8) for the well plate (100) (101).

6. Apparatus (1) according to any one of claims 1-5, wherein the pump (5) is submerged inside the tank (2) or located next to the tank (2).

7. Apparatus (1) according to claim 6, wherein the pump (5) is continuously operated.

8. Apparatus (1) according to any of claims 6-7, wherein the pump (5) is connected by a supply line (9) to an inner sub-tank (10) located inside the tank (2) ), and which in turn is directly connected to the needles (4).

9. Apparatus (1) according to claim 8, wherein the supply conduit (9) has an adjustable lateral exhaust path (1 1) that controls the flow of liquid exiting through the needles (4).

10. Apparatus (1) according to any of claims 1-9, further comprising a means (12) for heating the liquid contained in the reservoir (2).

eleven . Apparatus (1) according to claim 10, wherein the heating means (12) is a resistor.

12. Apparatus (1) according to any of claims 10-1 1, further comprising a thermostat (13) adjustable from outside the tank (2) to control the heating temperature.

13. Apparatus (1) according to any one of claims 1-12, wherein the liquid is oil.

14. Method for applying a liquid in at least one well (101) of a plate (100) with the plate (100) in an inverted position by means of an apparatus (1) according to any one of claims 1 -13 above, characterized in that it comprises the following steps:

placing a plate (100) in an inverted position on the needles (4) so that each of the needles (4) is inserted into one of the wells (101); Y

actuate the pump (5) to cause the liquid to rise through the supply line (9), reach the sub-tank (10) and ascend through the needles (4) until they exit through the tip of the same to be adhered to the wells (100) of the plate (100).

15. Method according to claim 14, further comprising the step of activating the resistor (12) to control the amount of liquid that is adhered to the wells (101) of the plate (100).

16. Method according to any of claims 14-15, wherein the liquid is oil.