CA1267083A - Sheet-like diagnostic device - Google Patents

Abstract

BEHRINGWERKE AKTIENGESELLSCHAFT 84/8 024 - Ma 518 Dr. Ha/Sd.

Abstract of the disclosure Sheet-like diagnostic device A solid diagnostic device for the quantitative determination of substances of biological affinity in biological fluids is described. A process is also described in which the biological fluid is brought into contact with a specific functional sector of the device, the fluid migrates through several functional sectors situated beside one another and containing suitable reagent components, and one or more substances of biological affinity are detected in such functional sectors which contain, for each substance to be detected, at least one combination partner of biological affinity, attached to a solid phase.

Description

67~

BEHRINGWERKE AKTIENGESELLSCHAFT 84/~ 024 - Ma 518 Dr. Ha/Sd Sheet-like dia~nostir device The invention relates to a solid diagnostic oevice ~hich comprises several functional sectors and is used for the detection and quantitative deterRIination of substances or analytes in biological fluids. The ;nvention also relates to a process using this clevice in ~hich, after the device has come into contact ~ith the fluid, the analytes react ~ith specific combination partners having biological affinity and are detected by means of labelling reagents.

In methods of diagnosis, the ability to identify and est;mate specific compounds has made it possible to mon-itor the administration of medicaments, the quantificationof physiologically active compounds or secondary products thereof and the diagnosis of infections. In this respect, the immunoassay methods tRIA, ELISA and the agglutination test) are of particular importance. The specific com-bination reactions utili~ed in the tests are not limitedto immunological interactions~ such as antigen-antibody or hapten-antibody ;nteractions, but also ut;li7e inter-actions having biological affinity, such as lectin-sugar or active compound-receptor.
Although the e~isting tests are sensitive and specific, they do not constitute convenient application forms, because of the long duration of the tests ~in most cases several hours or even days) and the frequent test stePs, such as immun~ reaction~ washing steps and enzymatic reaction. The long test times are not compatible uith use in emergency methods of diagnosis.

Integrated dry chemical test elements, such as are des-cribed in the present invention~ simplify the perfor-mance of the te!sts and shorten the test times.

~7~83

- 2 -No sheet-l;ke test element, ;n ~hich all the components of the immune reaction of a heterogeneous ilnmunoassay using solid phase detect;on, and the funct;onal per~ormance and the "bound-free" separation, are ;ntegratedh~s been des~r~d so fæ.~

~h~reas ;n the test strip assembly the immune reaction steps and the separation of bound and free phases are operated in the heterogeneous test by directed streams of l;qu;d, ;n test element assembl;es operat;ng by means of thin layers lam;nated over one another ~film tech-nology), processes controlled by d;ffus;on and d;rected by the concentrat;on gradient are~ possible driving forces.
A flu`orescence labell;ng ;s used in German Offenlegungs-schrift 30329,7Z8 (Japanese Patent P144,341/82) and EP A
OrO97~952 ~Japanese Patent 114,359/82)~ The labelling has a lo~ molecular ~eight and hence promotes processes controlled by diffusion. Houever, the test has to be carried out at an elevated temperature. In the first of these tuo cases both the free phase and also the bound phase are evaluated. In film technology the absorp~ion of solvent is effected either by hydrating s~ellable com-ponents or by f;lling capillary cav;ties. In the case of assemblies having layers laminated over one another only the top layer and the bottom layer are accessible to detection ~ithout major d;fficulties.
After the reaction steps have taken place it is difficult to react-reagents uith components in intermediately placed layers. In the test strip assembly having zones situated one behind another, such as is used in the present in-vention, i~ pr;nciple each zone ;s readily accessible,both from above and also from belo~, for a determination and also for the addition of reagents ~hich may perhaps be required.

~he invention relates to a sheet-like diagnostic devioe uhich contains all the reagent components and uhich con-tains not only ,3ll. the components required for the functional sequence, but also the functiônal sequences ;~267~83 themselves in an integrated form, and by means of which it is possible to detect an an~lyte having properties of biological aff~ity,-m such a way that a sc71ution of th~analyte is brought into contact with a functional region of the device designed for this purpose, and the analyte is detected via a signal-produc;ng system in a single functional region, a solid phase zone.

A second analyte, or further analytes, as constituents of the same solution can be detected at the same time by means of the device, if these analytes possess properties of biological affinity different from the first analyte.
They `are also detected in the same manner as the first analyte in a single functional region, a solid phase zone appropriate for them. The functional regions for the detection of the ~econd or further analytes are sieuated on the sheet~like device in front of or behind the functional region for the detection of the first analyte. The device can also contain several solid phase zones which are ap-propriate for an analyte and different measurement rangesof this analyte. The device contains all`reactants and reagents in a deh~drated form.

The sheet-like diagnostic t~vice comprises one or several strips, arranged behind one another, of material which have a capacity for absorbing aqueous solutions. The strips are fixed on a solid support . They contain the reagent components required for the particular diagnostic agent and thus become functional sectors or functional 3û regions. The functional sector situated at one end of the strip-shaped device (solvent application zone) is brought into contact with the analyte solution by being dipped into the latter or by the application of the latter. The solution migrates through all the functional regions.
The absorptive capacity of the supporting materials of which t~e strips are composed causes a flo~ of liquid which st~7s . at the other end of the str;p-shaped device.
The analyte can also be applied in the middle region of the device, and a flow of liquid from one end of the t~vice to the other can then be induced.

The sample does not have to be applied directly to the chromatographing section of the device ~ ~t can also be applied to an absorptive material which is situated on the ~evice and has the function of removing blood cells from the sample. After being filtered the sample then reaches the device - ~:. In the course of this fil-tration process the addition of reagents can be effected at the same time by dissolving the latter out of components present in the filter in a dry state. Interfering factors can be eliminated from the solution by means of such com-ponents. Thus, for instance, the ascorbic acid present in a sample, which interferes ;n the use of oxidases and peroxidases as labelling agents, can be rendered harmless by means of a suitable oxidizing agent. Furthermore, the filter can also have the function of an adsorbent wh;ch removes interfering factors from the sample by adsorption. The filtration, adsorption and reagent ad-mixing function for conditioning the sample for the testcan also be taken over by the mobile phase application zone or a zone situated behind the latter.

The distribution of the solvent in the individual function-al regions depends on the adsorpt;ve capacity and the dimensions of the materials used.

The solvent application zone can have the funstion of a volume metering element, as described in ~erman Patents

3,043,608 and 2,332,760, and US Patents 3,464,560, 3,600,306, 3,667,607, 3,~02,847, 4,144,306 and 4,258,001.
It can contain, in dry form, the various reagents required for the function of the test element. The solvent ap-plicat;on zone can be a piece of fabr;c paper which is located at one end of the test element and which becomes completely saturated with a definite volume of liquid merely by being dipped into a solution, for e~ample a solution of the sample, or by being briefly flushed with tap water, and then releases the liquid to ~he ~succeeding ;1~67~33 zones more slowly and ;n a controlled manner. The solvent application 20ne has dimensions such that it takes up sufficient liquid to permit the latt~r to migrate to the other end of the devlce, the end of the absorption zone.
s ~etween the solvent application zone and the absorption zone there are located the functional regions in ~hich are contained reaction components, for the performance of the test and in which all the reaction stages of the performance of the test take place. Part of the reaction components for the performance of the test can also be housed in the sample application zone. The absorption zone has he function of absorbing excess and freely mobile reagent components and reaction products of the signal-producing system.

The absorbent supporting materials in the form of one or more strips, as constituents of the various functional regions, can, according to choice~ be composed of cellulose, of chemical derivatives of cellulose or of plastics having a porous or fibrous structure and adequately hydrophilic properties, or of particles such as cellulose or silica gel embedded in a synthetic membrane, and also of natural products ~hich are hydrophilic but have been rendered insoluble in water. A combination of strips composed of different materials can be used. Suitable absorbent materials are selected on the basis of the requirements set for the particular diagnostic device.

Reactants with immunological.~inding pr~perties such as antigens, haptens or antiboaies are incorporated in various embodiments of the device.
- - - - In the event that glycoproteins or oligosaccharides which attach themselves to lectins are to be detected, one reactant having bio-logical affinity can be the specific lectin, while thesecond reactant having immunological affinity can be an antibody which is directed against a point of attachment on the analyte other than that of the lectin. In the event that microbial active compounds are to be detected, - 6 - ~ ~6~ 3 one comb;nation partner can be the receptor substance for the active compound, while the second combination partner can be an antibody which is directed against an-other point of attachment on the active compound.

One combination partnar having biological affinity be-comes attached during the progress of the reaction, or has already been attached to the supporting material in the functional region ~esigned ~For the detection of the analyte (solid phase zone). It is also called the solid phase combination partner. The other combination partner~s) are present in the supporting materials. They are pro-vided ~;th a labelling.

Amongst the various known possibil;ties of labelling, enzyme labelling is preferred. It requ;res chromogenic substrate systems or substrate systems which produce fluorescence or chemiluminescence. Chemiluminescence labelling represents a further example of a labelling which is only measured after the ad~ition of a reagent.
It is possible to measure either the~chemiluminescence itself or a fluorescence excited by the latter. In most cases fluorescence labelling is measured without the addition of a reagent being required. However, as in the use of certain rare earth chelates, it can also be desir-able to produce the fluorophore to be measured only as the result of adding a reagent, or to add a second fluorophore which becomes excited by the first or ~hich excites the first fluorophore. The fluorescence can be 3G measured at one point, as a function of time or as fluorescence polarization.

A reagent required for detection can be induced to react with the immune complex to be detected in various ways, after the separation stage. Part of the signal-producing system can be located in the solid phase zone. After the solid phase has been adequately washed, a reagent required to detect the labelling can be released at a retarded ~ rate in various embodiments in the heterogeneous ~llZ~ 3 ~ 7 ~
immunoassay with detection in the bound phase. The follo~-ing are possible examples:

The application of reagents by means of a stream of liquid arranged parallel to the main stream of liquid, but flo~ing more slowly and starting from the mobile phase reservoir and entering in front of the zone containing the labelled component. The parallel stream of liquid can be controlled by using an absorbent medium ~hich chromatographs more slowly, for example a paper which chromatographs suitably slowly or a paper ~hich is im-pregnated in places with "components temporarily blocking the way", such as, for example, polymers ~hich impart a high viscosity on passing into solution (for example polyvinyl alcohols or dextrans).

After the solid phase has been adequately washed ~= com-pletion of chromatography), the application of reagents can be effected by pressing down an element ~h;ch is a solid constituent of the test element. The "pressing down" can be effected mechanically or by removing dis-tance pieces by the action of a stream of liquid. For example, the mechanical pressing doln of an element con-taining the reagents can be effected by pressing down a flap or a piece of paper supported by distance pieces.
The lo~ering of an element containing the reagents by the action of the stream of liquid can be effected, for example, by laminating over one another the solid phase, a ~ater-soluble polymer and the reagent carrier (for example a suitably impregnated piece of paper).

A retarded introduction of reagents into the liquid stream can be effected using a microencapsulated reagent which only emerges from the encapsulation after the solid phase has been adequately washed, or by coating the reagent adhering in the matrix with components which dissolve s low ly .

7~83 One possible means presented for the special case of enzyme labelling is as follows: when a perox;dase labelling is used, a glucose oxidase zone can be placed in front of the solid phase zone. ~lucose and also the chromogen are then incorporated into the liquid stream, ~hich can result in color formation behind the glucose oxidase. Appreciable color formation is only observed if, at an appropriately high concentration of perox;dase, sufficient H2O2 is formed by the oxidase. This formation of the peroxide sets in slo~ly, reaches an optimum con-centration arld finally reaches a high concentration which results in inhibition of the enzyme and thus automatic cessation of the color formation. This coloration can be moderated if an H2o2-acceptor~ for example a thioether as a mild reducing agent, or the enzyme catalase is incorporated in the oxidase zone or in front of the latt~r.

In this example a reagent for detecting the labell;ng is 2~ produced by a delay circuit, making use of an enzyme.
The coLor formation in the solid phase zone only begins after this zone has been adequately ~ashed free from non-specifically bound labelling by the stream of Liquid.

There are several possible means of preparing the solid phase zone. The components fixed there can be attached by chemical covalent bonds or adsorptively to an absorptive support ~hich is a part of the test element.
These components can also be attached to a clispersion of particLes which remain fixed at the place of application after they have beenapplied to an absorbent support. For example, suspensions of cells carrying specific receptors on their surface, such as, for instance, Staphylococcus aureus Cowan I cells, or latex particles carrying com-bination partners of biological affinity attached totheir surface, are suitable for being fixed in a paper matrix. The components of the test strip ~hich are attached to pipettable supports and also the unattached ~Z~;7~83 _ 9 _ components of the device can be dried onto the absorbent matrix of the element by air drying; freeze-drying stages are not absolutely necessary.

A few test performances will be illustrated as examples of embodiments which can be regarded as independent of the labelling used. For the sake of simplicity, they are only described for the detection of a single analyte by means of the diagnostic device.

The following two embodiments, uhich conform to the principle of competitive immunoassay, will be described for the case where the analyte has only a single com-bination point of biological affinity or only one com-bination point of biological affinity out of several isutilized:

The solid phase combination partner is attached by covalent bonds or adsorptively to the supporting material of the solid phase functional region. The solution of analyte renders mobile a predetermined amount of labelled analyte contained in the diagnostic agent. The two com-ponents migrate into the functional sector containing the solid phase combination partner and compete for com-bination with the solid phase combination partner. If theproportion of analyte is high compared ~ith the labelled analyte, little labelled analyte ~ill be attached. If it is lo~, a great deal of labelled analyte will be attached.
The solid phase combination partner is housed as an un-attached component in a functional region in front of the solid phase functional region. The oncoming front of solvent transports it into the solid phase functional region, where it becomes attached. This solid phase attachment is produced by combination systems of bio-logical affinity which are independent of the combination system of the analyte. A combination partner which is conjugated with b;otin attaches itself to avidin attached ~2~7~33 to the support. An immunoglobulin, such as IgG, as a combination partner, is f;xed via its Fc component to support-attached protein A of S. aureus, or is attached by solid phase antibody of another species, nc~ idio~pically directed 5 to said i~nrn~oglobul~n.
As previously described, the analyte and the labelled analyte compete, as constituents of the di3gnost;c agent, for the attachments to the solid phase combination partner during the procressing period. This competition reaction takes place partly with the dissolved solid phase combination partner and partly with the solid phase combination partner which has already been attached to the s`olid phase.
If two combination points of differing specificity are present in an analyte, several embodiments, conforming to the principle of sandwich immunoassay, of the diagnostic agent are conceivable. Two of these will also be il-lustrated below:

If the solid phase combination partner is attached bycovalent bonds or adsorptively to the supporting material of the solid phase f~nctional region, the analyte forms, ~ith the labelled combination partner, a binary complex ~hich migrates together ~ith the solvent into the solid phase functional region and reacts there with the solid phase combination partner, ~ith the formation of a ternary complex, attached to the solid phase, which can be det-ected via the labelling of the first combination partner.The excess labelled combination partner is removed by the solvent into the subsequent functional region, the absorption zone.

If the solid phase combination partner is present in a non-attached form in the diagnostic agent and is rendered mobile by the solvent, the two reactants of the analyte of biological affinity are housed in the functional regions in such a ~ay that the analyte reacts simultaneously or successively with both reactants and the resulting ~Z~ 33 ternary complex then m;grates into the solid phase functional region, where, as already described above, it becomes attached to the sol;d phase via a second system of biological aff;nity which is independent of that of the analyte.

In order to illustrate the embodiments described above and further embodiments which conform to the immunometric test princ;ple, the pr;nciple of indirect antibody det-ection or the ELA ~en2yme-labelled-antigen) princtple of immunoassay, summary tables I and II i llustrate in an exemplary manner the distribut;on of the components of the agent in the functional regions and, after the performance ~f the reaction, the composition of the solid - 15 phase complex, the amount of which is a measure of the concentration of analytes in the sample.

~267~33 .. 1- ~'--X , ~ ." ~ ~ Q, o il ~_ E tl~ ~ x ~ O

C~
V~

1~76~83 ~ L

o ~: a ~ :D ~ (~ ~; 1 o vO) O ~ =O ~
_ ~ ~ ~ - ~ 1 ~ 1 8 ~0~~

l 1 ---1 ~ ~ ~ ~ 'V 0~
L .~ ~ aJ ~ D ~1 ~
L ~ ~ ,~ ~ C .~ C ~
_1 ~ Q 3 0 ~ . ~ C _ _ c ~ c ~ ~, ~
, x ~ x C ~ ~ D
m ~~ al v, ~ ~ o UJ _ S
V~

~ 2~ V83 It has been found that a completely integrated test strip operating in accordance ~ith the principle of heterogeneous immunoassay by means of solid phase detection is not only feasible in principle~ but can, in addition, also be evaluated with;n a period of less than one hour, the quantification and the sensitivity of conventional RIAs or ELISAs being achieved. The detection of trace com-ponents ;n the range of 10-12 mol~liter has been made possible at reaction times of less than 30 minutes, at room temperature, the amounts of sample required being ~ 6 mol, corresponding, for example, to approx. 1 pg.
The arrangements described also enable tests of lower sensitiv;ty requirements to be carried out, ho~ever.
Standard curves over two to three decades were obtained ~hen evaluation ~as carried out with the Sanoquell reflect-ometer (made by Quelle). The chromatography time for the test element, ;ncluding complete color development, is not more than 16 minutes. Evaluation can also be carried out visually. With HCG as analyte, the start of the range of determination in an example usirg a glucose oxidase attached to a solid phase and a peroxidase labelling ~as 0.3 ng/ml (corresponding to 3 U/liter).

In the example fGllowing, the application of the principle of the competitive double antibody test is presented as a concrete embodiment. In this test configurat;on, four components ha~e to be reacted successively for the det-ermination reaction and the separation stage, the re-action times and the concentrat;ons of the reactant being cr;tical values. The example is not to be regarded as limiting in any way, but merely serves to illustrate the subject of the invention further.

Example Completely inte!grated enzyme-;mmunochemical device for the detecticn of HCG by means of a bu;lt-in chromogen substrate system.

- 15 - ~ ~67~3 1.1. ~eagents 1.1.1. HCG-perod;xase conjugate HCG hav;ng a spec;f;c act;v;ty of approx. 3000 U/mg was - 5 obta;ned from Organon. Perox;dase from horserad;sh was obta;ned from Boehringer Mannheim (catalog no. 413,47û).
The hetero-bifunctional reagent N-~-male;m;dobutyryloxy-succinim;de t6MBS) was obta;ned from Behring D;agnost;cs and was-reacted ~;th the HCG as descr;bed by Tan;mori et al., 1983, ;n J.Imm. Meth. 62, 123-131. 2-;m;noth;olane hydrochloride tS;gma, catalog no. I 6256) was reacted with peroxidase as descr;bed by K;ng et al., 1978, ;n B;ochem;strx 17, 1499-150S. A conjugate was prepared from the GMBS-HCG and the iminothiolane-peroxidase as descr;bed by Tanimori et al. The crude conjugate was purified by gel chromatography over~Ultrogel ACA 44 (LK8).
The fraction in ~hich about 1-2 peroxidase molecules were coupled per HCG molecule was used for the test. The con-jugate was diluted uith Enzygnost IgE incubat;on med;um made by Behringwerke, order no. OS D, des;gnated br;efly as incubat;on med;um in the following text.

1.1.2. Antibod;es Antibodies aga;nst HCG were obta;ned by immunizing rabb;ts~
and antibodies against rabbit-IgG were obtained by immunizing goats. The IgG fractions ~ere isolated from serum by ammonium sulfate precipitation and an;on exchange chromatography, and were purif;ed further by ;mmunoadsorpt;on. The methods used are described ;n the book "Immunologische Arbeitsmethoden" tImmunolog;cal work;ng methods), Helmut Fr;emel, Editor, 1984, Gustav Fischer Verlag, Stuttgart. The anti-HCG antibody was finally diluted ;n the conjugate dilut;on buffer ;nd;cated above.

1.1.3. Glucose oxidase Glucose ox;dase from Asperg;llus n;ger was obta;ned as a ~ ~R~

~7~183 solution containing 300 U/mg (Serva, cataLog no. 22,737).
The glucose oxidase was finally diluted with incubation medium.

1.1.4. Glu~ose and tetramethylbenzidine c~-D-glucose and tetramethylbenzidine hydrochloride were obtained from Serva, catalog no. 2Z,720 and 35,926, respectively.

1.2. Preparation of the device The sheet-like functional regions were prepared as follows:

The mobile phase application zone was prepared by cutting, to dimensions o~ 20 x 6 mm, a fabr;c sponge cloth made by Kalle, this is a synthetic sponge of regenerated cellulose which has been compressed in a dry state. It ~as impregnated with a solution of 50 mg of glucose and 0.75 mg of tetramethylbenzidine hydrochloride per ml of ~ater, and was dried in a stream of air.

The conjugate, the anti-HCG antibody and glucose oxidase t5 ~i of each at 25 ~l/ml, 100 ~l/ml and 0.1 mg/ml~ res-?5 pectiveLy) ~ere applied behind one another~ at uniformdistance, to a 45 x 5 mm piece of MN no. 1 paper (Macherey & Nagel~, and were dried in the air.

A piece measuring 5 x 5 mm of Schleicher ~ Schull no.
597 paper was coated in a covalent manner with anti-rabbit IgG-antibody as the solid phase zone. This was effected by coupling the antibody with the paper, which had been activated with cyanogen bromide, as described by Clarke et al., 1979, Meth.Enzymology, volume 68, 441-442.
A Z0 x 5 mm piece of Schleicher ~ Sch~ll no. 2668/8 paper was used as the absorption zone.

- 17 - ~ ~6~3 The four pieces of paper, with a 0.5 - 1 mm overlap behind one another, were fixed on a ~lastic r~n by means of doubLe-sided adhesive tape ~Tesaband made by 3eiersdorf), so that a test strip 5 mm wide ~as formed.
1.3. Performance of the test The test ~as carried out in each case by applying 200 ~ul of an HCG dilution in incubation medium to the fabric.
1.4. Results The chromatographic development of the test element and the self-actuating color development ~ere complete after 15 minutes at room temperature, and evaluation could be carried out either visually or by means of a reflectometer.

The follo~ing values were obtained ~hen evaluating the solid phase zone (no. 597 paper) ~ith the Sanoquell blood glucose evaluation apparatus made by ~uelle:

HCG concentration ¦Measured values (U/liter) ¦(mg of glucose per dl of blood) . . . __ .
250.3 107 300 7û

3~
The following values were obtained with the same test strips using the Rap;mat urine test strip evaluation apparatus made by ~ehring~erke:

. .

7~8~

HCG concentration Measured values (U/liter) tBIT) 0.3 76 The test strip assembly shown here can also be achieved if the glucose oxidase and the anti-HCG antibody are located in the same zone. The ~es~ strip , which is cor-respondingly shorter, then ren~ers the result aft:er appr~x 10 minutes.

Claims (13)

THE EMBODIMENTS OF THE INVENTION IN WHICH AN EXCLUSIVE
PROPERTY OR PRIVILEGE IS CLAIMED ARE DEFINED AS FOLLOWS:

1. An analytical device for the detection or determination of a component in a fluid as an analyte with bioaffinity binding properties said device being composed of several sheet-like zones which are arranged behind one another and are in adsorbent contact with one another through their edges, containing a mobile phase application zone (MPAZ) at one end of the device and an adsorption zone (AZ) at the other end and also further adsorptive zones situated intermediately wherein reactants capable of interactions, of biological affinity, with the analyte are arranged in such a way that reactants capable of reacting with one another are present, separated spacially, wherein a) a reactant is fixed to a solid phase zone (SPZ) by means of covalent bonds or adsorptively or via an interaction of biological affinity in a zone which is located between the MPAZ and AZ and is in contact with the AZ, or becomes attached in a reaction which takes place in the device through a further reactant which is fixed in the SPZ by covalent bonds or adsorptively or via an interaction of biological affinity, b) a further labelled reactant (conjugate) is located, unattached, in a zone between the MPAZ
and the SPZ, and c) the analyte application zone is the MPAZ or a zone between MPAZ and AZ.

2. A device as claimed in claim 1 for the detection of two or more analytes each of which has one or more attachment points of biological affinity, which contains, per analyte, a spacially separated solid phase zone which is provided with combination partners attached to the support and specific for the particular analyte, and in which agent the analytes are detected separately.

3. A device as claimed in claim 1, wherein the MPAZ has the function of a volume metering element and releases to the subsequent zones at least sufficient liquid for the liquid, controlled by capillary forces, to reach the end of the AZ.

4. A device as claimed in claim 1, wherein the MPAZ is a plastic sponge or a particulate layer which is composed of hydrophilic polymers and which can, if appropriate, contain chemicals, buffer sub-stances or other substances required for the test.

5. A device as claimed in claim 1, wherein the sample application zone retains blood cells.

6. A device as claimed in claim 1, wherein the sample application zone is laminated onto one of the sheet-like zones of the chromatographing section of the device and is in adsorptive contact with this zone.

7. A device as claimed in claim 1, wherein all or some of the reagents required for the detection of the labelling are present in one or more of the sheet-like zones of the device or in a zone which is laminated onto one of the sheet-like zones of the chromatographing section of the device and is in adsorptive contact with this zone.

8. A process for the detection or determination of a component in a fluid wherein said component is an analyte with bioaffinity binding properties by rehydrating or solvating reactants and reagents by the fluid containing the analyte or by an additional fluid, said reactants and reagents being present in a dehydrated state in an analytical device for the detection or determination of a component in a fluid wherein said component is an analyte with bioaffinity binding properties, comprising a layer of a plurality of sheet-like zones adjacent one another and in absorbent contact with one another, said layer including:
a mobile phase application zone (MPAZ), an intermediate zone (IZ) and an adsorption zone (AZ), liquid being capable of moving by adsorption from said MPAZ
through said IZ to said AZ, and wherein said IZ further comprises a solid phase zone (SPZ) having at least one unlabelled reactant, capable of interactions of biological affinity with at least one analyte; or wherein said IZ is capable of having at least one unlabelled reactant fixed thereto which is capable of interactions of bioaffinity with at least one analyte, during analysis said at least one unlabelled reactant being fixed to at least one second reactant which is fixed to said solid phase zone;
at least one unattached labelled reactant (conjugate), capable of interactions of biological affinity with said at least one analyte, disposed in a zone between the MPAZ and the SPZ; and an analyte application zone disposed at said NPAZ or in between said MPAZ and said AZ;
said process comprising applying a sample to said analyte application zone, reacting the at least one analyte in the sample in said layer and detecting said at least one analyte in said layer.

9. The process as claimed in claim 8, wherein, after the liquid sample containing the analyte has been fed to the MPAZ or after the sample has been fed to a sample application zone and a mobile phase has been fed to the MPAZ, the liquid reaches the end of the AZ, under the control of capillary forces, and reactions between re-actants contained in the device and the analyte are there-by set in operation, and, after the labelling which is not attached to the solid phase has been removed chromatographically, the amount of the labelling in the solid phase zone, which is a measure of the analyte concentration in the sample, is determined.

10. The process as claimed in claim 8, wherein the reactions taking place in the device are based on the principles of immunological detection reactions, of com-petitive immunometric or sandwich immunoassay or of in-direct antibody detection by means of a labelled anti-body or of antibody detection by means of a labelled antigen.

11. The process as claimed in claim 8, wherein the labelling agent is a fluorophor which is detected or measured directly or is detected or measured after the addition of a reagent present in the device, or a fluorophor which is detected or measured directly or after the addition of a further reagent is formed from the labelling agent by the addition of a reagent present in the device.

12. The process as claimed in claim 8, wherein the labelling agent is a compound which can be excited to give chemiluminescene, it being possible to detect or measure chemiluminescence after the addition of a reagent present in the device.

13. The process as claimed in claim 8, wherein the labelling agent is an enzyme the activity of which is determined with the aid of a reagent present in the device.