COUNTING CHAMBER AND METHOD FOR MANUFACTURING A COUNTING CHAMBER

The invention relates to a counting chamber for determining the number of particles present in a fluid, said counting chamber substantially consisting of two parallel, at least partially transparent plates, which are bonded together by means of an adhesive and between which a practically entirely enclosed space is present, said space having a substantially constant dimension in the direction perpendicularly to said plates, which dimension will hereafter be called the cross dimension or the width of the space.

A counting chamber of this type is for example used for carrying out a semen analysis, whereby the sperm concentration of a sperm sample is determined. The small spacing between the surfaces of the glass plates leads to a capillary action, so that a sample charged to the counting chamber will spread over the space of the counting chamber. A very thin layer of the material of the sample is then formed, which can subsequently be observed optically, possibly by means of a microscope. When one of the glass plates is for example provided with a check pattern, it becomes possible to count the number of particles per unit area which are present in the sample.

The space of a counting chamber used for semen analysis has a cross dimension of for example 20 μm, whilst the space of a counting chamber used for analysing blood or other body fluids will have a cross dimension of for example 100 μm.

In order to obtain a counting chamber comprising a space which has a constant predetermined cross dimension, one of

the plates may be locally provided with a coating having a predetermined thickness, after which the two plates are bonded together at the location of said coating. In that case the space of the counting chamber will be bounded by the coating that has been applied and will have a cross dimension which corresponds with the thickness of the coating. Although it is possible in this manner to make a counting chamber comprising a space which has a very precise cross dimension, the manufacture of such a counting chamber is a relatively costly process which is difficult to control.

The object of the invention is to provide a counting chamber which can be manufactured in a simple and reliable manner.

In order to accomplish that objective the counting chamber is provided with plates whose facing sides are substantially flat, and the cross dimension of the space is substantially equal to the thickness of the adhesive layer present between the plates. This makes it possible to manufacture the counting chamber of flat plates, for example flat glass plates, whereby a particular cross dimension of the space can be obtained by using an adhesive layer of a particular thickness.

According to another aspect of the invention the adhesive contains granules having dimensions which determine the spacing between the plates. Preferably said granules are spherical and consist of glass or a similar material. The presence of the granules prevents the adhesive layer from being flattened during the bonding process.

According to another aspect of the invention the adhesive contains granules having a predetermined maximum diameter of approximately 10 - 150 μm, preferably approximately 20 ■ 100 μm, as well as granules having a smaller diameter. The thickness of the adhesive layer will substantially correspond with the dimension of the granules having the maximum diameter, whilst the granules having a smaller diameter are not a determining factor with regard to the cross dimension. In this manner a very accurate width dimension of the space, which is constant over the entire space, can be obtained in a reliable and simple manner.

According to another aspect of the invention a transparent part of at least one of said plates may be provided with a grid or a check pattern, for example consisting of squares. The number of particles per volume unit of the fluid present in the counting chamber can be determined in a simple manner by counting the number of particles present within one square.

The invention furthermore relates to a method for manufacturing a counting chamber, wherein two at least partially transparent plates are bonded together in mutually parallel relationship, whereby the thickness of the adhesive layer is according to the invention substantially equal to the cross dimension of the space of the counting chamber. The adhesive may thereby comprise granules having a dimension which determines the thickness of the adhesive layer.

According to another aspect of the invention the adhesive contains spheres having a predetermined maximum diameter of approximately 10 - 150 μm, as well as spheres having a smaller diameter, which spheres are selected by sifting

spheres of various diameters. A problem in the production of spheres, for example glass spheres, is to produce spheres having a predetermined average dimension (diameter) within a predetermined range of dimensions. When said spheres are sifted mechanically by means of a screen which is set to a diameter which approximately corresponds with the average diameter of the spheres, this results in an amount of spheres, a relatively large number of which corresponds with the diameter to which the screen is set. The spheres selected by sifting may then be added to the adhesive with which the plates of the counting chamber are bonded together.

According to another aspect of the invention the granules are sifted prior to being added to the adhesive, by means of a screen having gaps of a predetermined width. Since gaps having a predetermined width are used, only those granules which are capable of being incorporated in an adhesive layer which corresponds with the width of the gaps can pass said gaps. The sifting of the granules preferably takes place in a fluid, for example water, so that the granules can be transported over and/or through the screen in a simple manner.

In order to prevent the granules present in the adhesive from staying together, thus forming lumps, the granules may be electrically charged in such manner as to cause mutual repulsion among said granules. When glass granules are used, this may for example take place by rubbing the granules in a woollen cloth.

According to another aspect of the invention the lumping of the granules is prevented by providing the granules with a

coating of a material whose cohesion is smaller than the adhesion between said adhesive and said material.

According to another aspect of the invention a mould containing a material having substantially the shape and dimension of the space of the counting chamber is placed on a flat side of one ot said plates, after which an adhesive is applied, which adhesive is skimmed off by means of an object having a substantially straight edge, which is moved over said mould. As a result of this the adhesive will be applied to the places present between the material of the mould, in a thickness which is determined by the thickness of the material of the mould. This makes it possible to apply a properly distributed adhesive layer of a predetermined thickness in an accurate manner. Preferably said thickness is slightly greater, for example 1 - 10%, than the cross dimension of the space of the counting chamber and the maximum dimension of- the granules present in the adhesive, so that it will be possible to exert sufficient force on the adhesive when pressing down the adhesive between the plates.

According to another aspect of the invention the mould consists of a plate-shaped element, which comprises unperforated material at the location where the space of the counting chamber is to be formed, and which is perforated in those places where the adhesive is to be applied. Such a mould may be obtained by locally providing a layer consisting of a gauze or an apertured fabric with an impermeable material, which bonds in said layer, so that the adhesive, when being skimmed off, can only pass through the layer at those locations where no impermeable material is present. A similar technique is used in screen-printing.

According to another aspect of the invention an adhesive is used which slightly shrinks during the curing process, so that the plates are slightly pulled down on the granules during said curing.

Further aspects of the invention, which may be used separately as well as in combination with each other, will be explained in more detail in the description of the Figures and be defined in the claims.

Hereafter an embodiment of the invention will be described by way of illustration with reference to the drawing.

Figure 1 is a front view of a counting chamber formed by two bonded-together glass plates;

Figure 2 is a plan view of the device according to Figure

1;

Figure 3 is a sectional view along line III-III in Figure

1; and Figure 4 is a sectional view of two bonded-together glass plates .

The Figures are merely diagrammatic illustrations of the embodiment, for the sake of clarity the dimensions are not shown in their actual proportions. Like parts are numbered alike in the various Figures.

Figures 1, 2 and 3 shows two glass plates 1, 2, which are bonded together by means of an adhesive layer 3. The adhesive layer 3 is not present on the entire surface of the glass plates 1, 2, but only in circumferential area 4, as is shown in Figure 1. A space not containing any adhesive is present within circumferential area 4, so that

a chamber 5 is formed between glass plates 1, 2. Said chamber 5 communicates with the outside via an opening 6.

In this manner a counting chamber for single use has been formed, which may have a width of for example 10 - 20 μm between the substantially parallel glass plates. When a fluid is charged to the chamber 5, for example by means of a pipette, said fluid will spread over the entire chamber, at least over a large part thereof, as a result of the capillary action, so that a thin layer of fluid is formed, which has a predetermined thickness everywhere. Then this fluid may be observed, for example by means of a microscope, in order to be able to detect particles which may be present in said fluid. As already said before, a semen analysis may be carried out in such a chamber.

The two glass plates may be entirely transparent, but it is also possible to provide a check pattern on one glass plate or on both, in order to make it easier to count the number of particles per unit area present in the fluid.

It will be apparent that a suitable counting chamber 5 can only be formed when the adhesive layer 3 has a constant thickness over the entire surface on which the adhesive layer is present. Because the adhesive layer is very thin, for example between about 10 μm and 100 μm, and because a high degree of accuracy of the thickness of the adhesive layer is required, it is very difficult to manufacture the illustrated device by maintaining the required spacing between the glass plates by mechanical means during the curing of the adhesive.

Figure 4 diagrammatically shows on a slightly enlarged scale the manner in which the thickness of the adhesive

layer is maintained at a predetermined level. To that end the adhesive layer 3 present between glass plates 1, 2 comprises spherical glass granules 7, 8, which have been selected such that the diameter of the largest granules 7 corresponds with the thickness of the desired thickness of the adhesive layer. Smaller granules 8, which may also be present in the adhesive layer, do not have an influence on the thickness of the adhesive layer. Since the granules have been selected by means of a mechanical screen, it is possible that said smaller granules 8 are contained in the adhesive layer, but the adhesive layer thickness is determined by the largest granules 7 that were able to pass said screen.

In practice it has become apparent, that it is possible in a simple manner to achieve a constant predetermined thickness of the adhesive layer by using spherical glass granules in the adhesive, which glass granules have been sifted by mechanical means.

The adhesive used may be a silicone adhesive, preferably an adhesive is used which contains acrylic monomers and/or acrylic urethane oligomers. An adhesive of this type is commercially available under the name "Viprocoll Adhesive UV2000" or "UV200 Stabifix Adhesive" from Visprox B.V., Haarlem (ΝL) , for example.