FI78180C - REAKTIONSKAERL FOER IMMUNOLOGISKA BESTAEMNINGAR. - Google Patents

Description

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Reaction vessel for immunological determinations Reaktionskärl för immunologiska bestämningar

The invention relates to a tubular reaction vessel for use in immunological assays, to which an antibody to which the substance to be measured (antigen) and the tracer (labeled antigen) bind is attached to the inner wall of the closed or lower end of the vessel.

Reaction vessels made of a test tube by coating the inner wall of the tube with an antibody are already known. Systems are also known in which an antibody-coated sphere or other shaped body to which a test substance and label binds is dropped into a conventional untreated test tube (US 4,320,087, 4,225,575).

The disadvantage of the above reaction vessels is the long binding time required for the binding of the test antigen and marker to the antibody or to the antibody-coated bead or other body in the test tube. This is because the rate of the binding reaction is determined by the diffusion of the substance to be measured (antigen) and the tracer (labeled antigen) in solution in the tube to the walls of the tube. In a standard test tube, diffusion distances are long and binding is slow. As the size of a molecule increases, its diffusion rate decreases, so that, for example, the determination of high molecular weight protein hormones is slow. When using an antibody-coated bead, the rate of binding does not significantly increase compared to the coated tube. Dropping a sphere or other object into a test tube causes the assay performer one more step. The handling and storage of separate parts for the same reaction vessel system at 2,781,80 ti is also cumbersome, and the antibody-coated surface of the body to be dropped into the tube can be easily damaged, which can cause inaccuracies in the test results.

The test tubes or spheres and other bodies used as reaction vessels are generally made of polystyrene or polypropylene. The antibody is attached to their surface by adsorption or chemically through several steps. This type of fastening is complicated and it is very difficult to coat hundreds of thousands of tubes in equal parts in one or more batches.

The reaction vessel according to the invention avoids the above-mentioned drawbacks and is characterized in that the surface of the inner wall at or near the closed end of the reaction vessel is enlarged by one or more surfaces parallel to the longitudinal axis of the vessel.

According to one embodiment of the invention, the container is corrugated. Alternatively, the vessel is divided into several parts by partitions so that the liquid can flow freely from one part to another. The reaction vessel according to the invention may also comprise a tube inside which there is another smaller radius coaxial tube directed upwards from the bottom.

According to the invention, the lower or closed part of the reaction vessel is provided with at least one and suitably at most eight corrugations, which are preferably arranged symmetrically along the circumference of the tube. According to a preferred embodiment 3 78180, the container is provided with 4-6 symmetrically arranged corrugations. Each crimp may extend from the wall of the reaction vessel in a distance of about 1/8 - somewhat less than the half, such as 1/8 to about 5/12, preferably about 1/5 -1/3 of the container cross-section to reduce the inner diameter of the diffuusiomatkojen but still sufficient free liquid to allow flow inside the vessel. The longitudinal shape of the folds, on the other hand, facilitates easy and accurate emptying of the container. The corrugations run longitudinally along the reaction vessel for a distance sufficient to extend above the surface of the sample in the vessel. For example, this length generally corresponds to about 1/4 to 1/2, especially about 1/3 of the length of the vessel when the vessel is in the form of a microtest tube.

According to another embodiment, the reaction vessel according to the invention may be provided with partitions projecting inside the vessel and allowing the free flow of liquid between different parts of the vessel. For example, the partitions may form a substantially right angle to the wall of the container or may be brush-shaped. The above in connection with the corrugated container with respect to the number of partitions or ridges, their length in the longitudinal direction and their projection inside the container also applies in connection with this embodiment.

A very advantageous reaction vessel has proven to be a test tube with a lower or closed end provided with five brushes made symmetrically in the inner wall of the tube, projecting from the wall into the tube by a distance of 4,781,180 corresponding to about 1/3 of the cross-sectional diameter of the tube.

According to a third embodiment of the invention, the reaction vessel comprises two coaxial tubes of different radii, the inner tube projecting upwards from the bottom of the outer tube, thus limiting the annular space between the tubes. The inner tube can be formed by pressing the bottom of the outer tube upwards towards the opening. Alternatively, the inner tube may be attached to the inner surface of the bottom of the outer tube. The width of the annular space may vary, but is suitably about 1/5 to 1/3 of the inner diameter of the outer tube. One or both of the coaxial tubes may also be provided with corrugations or ridges or partitions projecting into the annular space, as described above. In this case, the inner and outer tubes are preferably provided, e.g., with the same number of interlocking corrugations (Fig. 4).

The reaction vessel of the invention is preferably made of a plastic or other material containing hydrophilic functional groups such as -COOH, -OH, -CHO, -NH 2, etc. Suitable plastics for use in the reaction vessel include, e.g., polyacrylate-ethylene copolymers; polystyrene derivatives such as polyaminostyrene, polystyrene acrylonitrile or polyvinyl benzyl chloride; polyhydroxyethyl methacrylate, tris-acrylic-NH2 ·

Experiments have shown that 5 78180 of the test substance (antigen) and the labeled substance (labeled antigen) from the solution in the reaction vessel of the invention rapidly bind to the antibody attached to the wall of the vessel. With the reaction vessel according to the invention, the immunological assay can be performed quickly and the possibility of error is reduced because the binding reaction can be almost completed before pouring the solution from the vessel and measuring the amount of tracer bound to the wall. This is a significant benefit to the user when you usually have to do hundreds of configurations a day.

The attachment of the antibody to the reaction vessel of the invention is simple and reliable because the material of the reaction vessel contains functional groups.

The accompanying drawing illustrates a reaction vessel according to the invention.

Figure 1 shows a longitudinal and cross-sectional view of a corrugated reaction vessel at the lower end.

Figure 2 shows a longitudinal and cross-sectional view of a reaction vessel divided into sections by vertical walls.

Figure 3 shows a vertical and cross-section of a reaction vessel formed of two coaxial tubes of different radii.

Figure 4 shows a cross-section of a reaction vessel in which the inner and outer racks of Figure 3 are corrugated.

6,78180

Figure 5 shows the percent binding of test substance (antigen) and tracer (labeled antigen) as a function of time as measured in a reaction vessel according to the invention (Graph A) and as measured in a standard test tube (Graph B).

As shown in Figure 1, the wall area of the reaction vessel has been increased by means of four corrugations P extending a short distance along the tube. In this way, the liquid-filled space V of the tube is made narrower. In this embodiment, the distance between two opposing corrugations is somewhat more than half the diameter of the pipe. The reaction vessel of Figure 2 has similar properties, in which the liquid space V is divided into four compartments by partitions or ridges S projecting into the tube by a distance of about one third of the diameter. According to Figure 3, the liquid space V is left between two coaxial and different radius tubes T1 and T2. As shown in Figure 4, the fluid space is between two coaxial tubes, as in Figure 3, but the outer tube X and the inner tube Y are each corrugated as in Figure 1.

The reaction vessel according to the invention is used in the following manner. The sample and tracer are dispensed into the reaction vessel. The sample antigen and the labeled antigen then bind to the antibody on the wall of the reaction vessel. After the binding reaction has progressed far enough, the solution is poured out of the reaction vessel and the amount of tracer adhering to the wall of the reaction vessel is measured. The amount of tracer can also be measured from the solution removed from the reaction vessel. The measurement method depends on the tracer used. Attachment of the antibody to the wall of the reaction vessel occurs by chemically binding the antibody to the functional groups contained in the material of the reaction vessel.

When prepared in this way, similar reaction vessels are obtained even though the batches are large and, in addition, they remain well in stock. Due to the shape of the reaction vessel, the binding of antigen and marker to the antibody attached to the wall of the reaction vessel is rapid. The reaction vessel can be used to determine all substances and combinations of substances that are immunologically detectable. The "antibody" attached to the wall of the reaction vessel may also be an antigen.

Claims (10)

1. Tubular reaction vessel for immunological determinations, to which the inner wall of the vessel, at least at its closed end or near it, is fixed an antibody, to which antibody the substance to be determined and the labeling agent binds from the solution in the reaction vessel, characterized in that the surface of the inner wall at the closed end of the reaction vessel or in the vicinity of it is enlarged by one or more surfaces parallel to the longitudinal axis of the vessel, which extend in one piece with the wall of the vessel and allow a free flow of the liquid inside the vessel. 2. A vessel according to claim 1, characterized in that the surfaces form radially aligned folds in the wall of the vessel. Vessels according to Claim 1, characterized in that the surfaces form radially inwardly extending axes or partitions formed in the inner wall of the vessel. Vessels according to claim 1, characterized in that the surface is formed by pressing the bottom of the vessel against its opening over a portion corresponding mainly to the reaction zone. 5. A vessel according to claim 1, characterized in that the surface is formed by attaching to the bottom of the vessel a coaxial tube with a smaller radius. Vessels according to claim 4 or 5, characterized in that one or both tubes are provided with radial creases or axes or partitions extending into the annular space between the tubes and which are preferably arranged overlapping relative to the tubes. each other. 7. A vessel according to claim 2 or 3, characterized in that the vessel is provided with a very tight, preferably four - six folds, aces or partitions. 8. A vessel according to claim 7, characterized in that the creases, the axes or the partitions extend