MXPA04012232A

MXPA04012232A - Hts-capable method and testing system for determining the interaction between a c-reactive protein and constituents that bind to a c-reactive protein.

Info

Publication number: MXPA04012232A
Application number: MXPA04012232A
Authority: MX
Inventors: Pommereau Antje
Original assignee: Aventis Pharma Gmbh
Priority date: 2002-06-12
Filing date: 2003-06-10
Publication date: 2005-02-25
Also published as: AR040259A1; BR0311709A; CN1745300A; WO2003107000A3; TW200411179A; DE10226011A1; ZA200409199B; AU2003250344A1; CA2489021A1; JP2005529348A; EP1516187A2; WO2003107000A2; RU2005100042A; NO20050037L

Abstract

The invention relates to an HTS-capable method and testing system for determining the interaction between a C-reactive protein (CRP) or C1q and constituents that bind to a CRP or C1q in order to determine the concentration of a solution containing CRP or C1q and to identify substances that influence the interaction between CRP or C1q and constituents that bind thereto, particularly the interaction between CRP and C1q.

Description

METHOD WITH HTS CAPABILITY AND TEST SYSTEM FOR THE DETERMINATION OF THE INTERACTION BETWEEN THE C-REACTIVE PROTEIN AND THE COMPONENTS UNITED TO THE C-REACTIVE PROTEIN The present invention relates to a method with HTS capacity and to a test system for the determination of the interaction between C-reactive protein (CRP) or C1q and the components bound to CRP or C1q, for the determination of the concentration of a solution containing CRP-o C1q, as well as for the determination of substances that influence the interaction of CRP or C1q and the components bound to them. C-reactive protein (CRP) is an acute phase plasma protein whose concentration in the serum increases rapidly and to a large extent after an infection or tissue injury (Volanakis (2001), Molecular Immunology 38, 189- 197). In the presence of Ca2 + ions, CRP binds to phosphoco-lina (PCh). The latter is widespread in the polysaccharides of pathogenic organisms and in membranes of damaged and necrotic cells. CRP bound to PCh can activate the classical complement cascade by binding to the C1q protein. The complement is part of the immune system and participates mainly in the immune defense that induces the antibodies. The three physiological functions of complement are the defense of bacterial infections, the union of congenital and acquired immunity and the elimination of immunocomplexes and apoptotic cells. The cascades of classical complement, alternative and mannose-lectin can be differentiated (Walport (2001), N Engl J Med 344, 1058-1066) The classic complement cascade leads to the lysis of bacterial cells and begins with the association of C1q to the antibody bound to the surface of the cell or to the CRP bound to PCh. The C-reactive protein is used as a marker and, in addition, as a predictor of coronary heart diseases, the most frequent cause of death in industrialized countries. (Rifai and Ridker (2001), Clinical Chemistry 47, 403-411) Under new research (Jialal et al (2001), Circulation 103, 1933-1935) it is accepted that a decrease in the level of CRP or A blockade of the effects of the effector induced by CRP, for example, of the activation of the complement through the binding to C1q, can be useful to prevent and treat coronary heart diseases. , for besides, a method that allows to easily determine the concentration of CRP and / or C1q in the blood plasma and that also allows to identify substances that act by modulating - activating or inhibiting - the interaction of CRP and / or C1q with other components , especially the interaction between CRP and C1q. Methods for determining the binding of CRP and C1q have been described, for example, by Jiang et al. (1991), J. Immunol. 146, 2324-2330 and Agrawal et al. (2001), J. Immunol. 166, 3998-4004. In both publications an ELISA method is used. In this case, it is a method that is very expensive in time and material, of many stages and not homogeneous. Thus, for the execution of the method, one of the reactants is always immobilized first in a solid phase. In addition, numerous washing steps are necessary. Lebdue et al. (1998), Ann Clin Biochem 35, 745-753 describe a method for the immunonephelometric detection of CRP. For this, the sample is incubated with antibodies against CRP, which have been loaded with polystyrene particles. A decisive drawback of this method is that in order to carry it out it is necessary to have a nephelometer and that a relatively large amount of sample has to be used. The mission of the present invention, therefore, was to provide a method that allows to determine in a simple way the concentration of CRP or C1q in the blood plasma and that, in addition, allows to identify substances, which act by modulating, activating or inhibiting the interaction of CRP or C1q with other components, especially the interaction between C1q and CRP, the method advantageously being distinguished from the state of the art, in the sense that it is more sensitive, more economical, saves material, is simpler and / or faster to perform than the methods described up to now. The mission is solved by a method for the determination of a binding event of two components linked to CRP or C1q, directly or through one or more components, comprising the following steps: (a) initially introducing a solution containing CRP or C1q, (b) adding to the solution containing CRP or C1 q at least one donor component or a group of components, containing at least one donor component, in the case of the donor component of a component comprising at least a compound or a group of compounds, which can emit a signal after excitation by a light source (donor group) and the donor component being able to bind to CRP or C1q, directly or through one or more components of the group mentioned component, (c) adding to the solution containing CRP or C1q at least one acceptor component or a group of components, which contains at least one acceptor component, treating them e in the case of the acceptor component of a component, comprising at least one compound or group of compounds, which capture the signal emitted by the compound or group of compounds according to (b) and can yield it in the form of electromagnetic radiation. ca (acceptor group) and the acceptor component being able to bind to CRP or C1q, directly or through one or more components of said group of components, (d) excite at least one compound or group of compounds according to (b) ) (donor group) by means of a light source, (e) detecting the electromagnetic radiation emitted by the at least one compound or group of compounds according to (c) (acceptor group) for the detection of the binding event. As regards the electromagnetic radiation emitted according to (c), it is preferably in this case fluorescent radiation. As for the light source according to (d), it can be, for example, a laser or a lamp, for example a helium or halogen lamp. The detection of the electromagnetic radiation according to (e) can be carried out, for example, with the aid of a photomultiplier. The components to be used according to the invention are or preferably comprise polypeptides. In a particularly preferred embodiment of the invention, at least one of the components is an antibody that can be a monoclonal or polyclonal antibody. In the case of using a group of components, they are preferably able to join spatially consecutive to CRP or C1q or to each other. In this case, the binding is carried out in the form of a binding cascade, as is done, for example, by the consecutive binding of antibodies (primary antibodies, secondary antibodies, etc.). In the case of the components directly linked to CRP or C1q according to (b) and according to (c), it is preferably in this case components that are bound in each case to a specific binding region or to a certain epitope of CRP or C1q, the CRP or C1q-linked component binding according to (b) to another binding site different from that of the CRP or C1q-linked component according to (c). In a further preferred embodiment of the method according to the invention, in the case of a component directly linked to CRP or C1q, it is a naturally occurring participant in the binding of CRP or C1q. In the case of CRP, this is especially preferred for C1q and vice versa. In the event that the donor or acceptor component is directly linked to CRP or C1q, it is in this case correspondingly of C1q or CRP, in the case of the donor or acceptor component that includes a donor or acceptor group. In the case of using a group of components that can form a cascade, it is treated in the case of C1q or CRP correspondingly of the first component linked to CRP or C1q, which can be linked to CRP or C1q through the donor or acceptor component. In the case of the remaining components of the group of components, antibody molecules are especially preferred.

If, for example, a solution containing CRP is available, then preferably the donor or acceptor component, including C1q, or the donor or acceptor component that binds CRP through C1q is encompassed in this embodiment. The donor or acceptor component can then encompass an anti-C1q antibody or a secondary antibody, which can bind anti-C1q antibody. Correspondingly, the other respective component (acceptor or donor component) encompasses an anti-CRP antibody or an antibody that can bind anti-CRP antibody or a ter-ciary antibody directed against the latter.

In a preferred embodiment, the at least one compound or group of compounds according to (b) (donor group) and / or according to (c) (acceptor group) are located on the particles, the average diameter being found of the particles preferably between 150 and 250 nm, particularly preferably at approximately 200 nm. The particles are then correspondingly encompassed by the donor and acceptor components. The particles can be manufactured in this case from a polymeric material. In the case of the method according to the invention, it is a method with HTS capability (high performance tracking), which can be carried out homogeneously in a one-stage process. In a preferred embodiment the method is carried out according to the "mix and measure" principle, which leads to a considerable saving of time and a greater precision of the measurement values. The method according to the invention allows the measurement of the binding of CRP to C1q and, thereby, the identification of substances that influence this binding, as well as the measurement of the concentration of CRP or C1q in complex media in less than one 10 percent of the time, which would have to be used to employ the inhomogeneous methods described so far. A special feature of the new method is that it allows to monitor the binding between the two proteins in solution, since no reactant has to be in-mobilized to a solid phase. The measurement can also be carried out directly with the proteins obtained from biological sources, without having to make a modification of CRP or C1q, for example by binding a fluorophore. A special advantage of the method is, therefore, that the protein can be tested in the native state and that the danger of denaturation, which would exist in the case of an immobilization and / or modification of the protein, is eliminated. A further advantage of the method according to the invention is that the sample volume of the method according to the invention is considerably reduced compared to traditional methods. Thus, volumes as small as 10 μ ?, can be used, which minimizes the waste of material. In addition, highly diluted solutions with CRP or C1q concentrations of less than 1 nM, even less than 100 pM, can be used. It is, therefore, a very sensitive method. In addition, based on the above, it is the first method that makes possible a high performance screening of substances, which affect the activation of the complement through the binding of CRP to C1q. In a preferred embodiment of the method according to the invention, the transmission of the signal of at least one compound or group of compounds according to (a) (donor group) at least one compound or group of compounds according to ( b) (acceptor group) takes place through an oxygen-singlet. In this case the donor group preferably comprises a compound, which can convert an oxygen triplet into a singlet oxygen after excitation by a laser and the acceptor group encompasses at least a first compound that can be excited by an oxygen-singlet, and a second compound that absorbs without energy the energy absorbed by the group - Si ¬ It is excited by the oxygen-singlet and can emit it in the form of fluorescent radiation. The formed oxygen-singlet can diffuse from the donor group to the acceptor group and can react with the chemiluminescent substance found there. The energy that is released with it is transferred to the fluorophores, which radiate energy in the form of fluorescent radiation, which can be detected with a photomultiplier. The premise for a detectable signal is the spatial proximity of donor and acceptor granules, since the singlet oxygen is unstable and decomposes into an aqueous solution. The binding components to be used by this method are selected, therefore in such a way, that at the start of the binding event, the distance between the donor and acceptor group is preferably less than 200 nm, since larger distances are not possible an effective energy transfer by oxygen-singlet due to the decomposition time of the oxygen-singlet. Particularly preferred in this embodiment are donor groups and acceptor groups located on particles, the particles preferably having an average diameter between 150 and 250 nm, particularly preferably approximately 200 nm. In this case, the particles are preferably used so that the particles containing the donor groups are adjusted to a final concentration of 1-40 μg ml and the particles containing the acceptor groups are adjusted to a final concentration of 1-80 μg. ? /? t ?? The binding capacity of the particles can be, for example, about 0.1 nM to 1 nM per g / ml particle. For the method, Alpha-Screen granules from Perkln-Elmer Life Sciences are used as particles, particularly preferably. In the case of this embodiment, the excitation of the donor group is effected by irradiation with a laser at a wavelength of 680 nm, and the radiation emitted through the acceptor group can be detected at a wavelength between 520 and 600 nm. The method can be carried out in a corresponding manner such that the donor and acceptor granules, which carry the donor and acceptor groups, are arranged. Now, the components bind to the donor and acceptor granules, which are able to bind CRP or Cq1, by themselves or through other components. As already mentioned above, these components are C1q or CRP, or an anti-CRP or anti-C1q antibody, or a secondary antibody directed against anti-CRP or anti-C1q antibodies. In a further preferred embodiment of the method according to the invention, the signal transfer of at least one compound or group of compounds according to (b) (donor group) is effected to the at least one compound or group of compounds of according to (c) (acceptor group) through the transfer of energy without emission, especially preferably through fluorescence resonance energy transfer.

In this case the method can be carried out, for example, in such a way that at least one compound or group of compounds according to (a) (donor group) comprises a compound containing a europium salt and the at least one compound or group of compounds according to 5 (b) (acceptor group) encompasses allophycocyanin (Grepin et al. (2000), Drug Discovery Today 5, 212). Alternatively, the donor or acceptor group is dyes that are suitable for energy transfer without emission. All those compounds known to those skilled in the art, which are suitable for the transfer of energy without emission, especially for the transfer of energy by fluorescence resonance can also be used (see, for example, Pope et al. 1999), Drug Discovery Today 4, 350). The binding components to be used in this embodiment are chosen, preferably in such a way that the distance between the donor and acceptor groups after bonding is less than 10 nm, since at a greater distance an energy transfer is not possible. effective without issuance. The method according to the invention is used, in a preferred embodiment according to the invention, for the determination of 0 the concentration of a solution containing CRP or C1q, of unknown CRP or C1q content, in which The solution is preferably blood plasma or blood serum, or blood plasma or blood serum diluted with an appropriate physiological buffer or with water. This method can be used for diagnostic purposes, especially to determine the state of inflammation of an organism and / or to determine the risk of heart infarction and / or stroke. The object of the present invention is therefore also a diagnostic test system with HTS capability, to determine a binding event between CRP or C1q and a component linked to CRP or C1q, which comprises the following components: (a) at least one donor component or a group of components containing at least one donor component, wherein the donor component of a component comprising at least one compound or group of compounds which, after excitation by a light source, is treated; can emit a signal (donor group), and the donor component being able to join CRP or C1q, directly or through one or more components of the mentioned group of components, (b) at least one acceptor component or a group of components, which contains at least one acceptor component, wherein the acceptor component of a component comprising at least one compound or group of compounds that captures the signal emitted by the The component or group of compounds according to (b) can be assigned in the form of electromagnetic radiation and, the acceptor component being able to bind to CRP or C1q, directly or through one or more components of said group of components. Said assay system with HTS capacity comprises in this case, in addition, the following components: (c) blood serum or blood plasma or blood diluted with a physiological buffer or by water, (d) a light source for the excitation of the at least one compound or group of compounds according to (a), (e) a detection system for detecting the electromagnetic radiation emitted according to (b). In another preferred embodiment of the method according to the invention, at least one test substance is added before step - (a) and / or before step (b) and / or before step (c) and / or before stage (d), in order to observe if this has an influence on the binding event. In this way substances can be determined, which act by modulating, inhibiting or activating the interaction between CRP or C1q and a binder, especially the interaction between CRP or C1q. Preferably, for this a database of test substances in HTS processes is tracked, in order to determine a substance with the desired properties.

Object of the present invention is, therefore, also a test system with HTS capability for the determination of active substances, which act by modulating the interaction between CRP or C1q and a component linked to CRP or C1q, which comprises the following components: (a) at least one donor component or a group of components containing at least one donor component, wherein the donor component of a component comprising at least one compound or group of compounds which, after excitation by a source, is treated of light, can emit a signal (donor group), and in which the donor component is able to join CRP or C1q, directly or through one or more components of the mentioned group of com ponents, (b) so less an acceptor component or a group of components, which contains at least one acceptor component, wherein the acceptor component of a component comprising at least one compound or group of compounds that receives the signal emitted by the compound or group of compounds of according to (b), and may assign it in the form of electromagnetic radiation, and in which the acceptor component is able to bind to CRP or C1q, directly or through one or more components of the mentioned group of components, (c) at least one test compound. Said test system with HTS capacity preferably comprises in this case the following components: (d) a light source for the excitation of the at least one compound or group of compounds according to (a), (e) a detection system for detect electro-magnetic radiation emitted according to (b), (f) a solution containing CRP or C1q. In this case, the MTS-capable test systems according to the invention are preferably used for carrying out the method according to the invention described above. Special forms of embodiment of the ingredients and components of the test systems correspond, therefore, to the special embodiments mentioned above, which are used in the method according to the invention. In Diagram 1 the method described in the exemplary embodiment is schematically represented. In this case an anti-C1q antibody is used as a donor component, the donor group being located on particles that are bound to anti-C1q antibody. The donor component can be linked in the present case to CRP through C1q. In the case of the acceptor component, in the present case, it is a secondary anti-rabbit antibody, the acceptor group being located on particles that are linked to the secondary anti-rabbit antibody. The acceptor component can be linked in the present case to CRP through a rabbit anti-CRP antibody.

Example of embodiment: binding assay for the determination of the binding of CRP to Clq To carry out the assay the Alpha-Screen detection kit of Packard Bioscience is used. This kit comprises donor and acceptor granules, comprising compound donor granules which, after excitation by a defined wavelength of triplet oxygen, can be converted into singlet oxygen, and comprising compound acceptor granules which they can be excited by oxygen-singlet, as well as compounds that accept the energy of the excited compounds in the form of energy transfer without emission and can then emit in the form of fluorescent radiations. The mentioned compounds are embedded in this case respectively in a hydrogel matrix. The granules used were previously coated by the manufacturer, namely the streptavidin donor granules and the acceptor granules with anti-rabbit antibodies.

Table 1: Overview of the materials used Reagents / Plates Molecular weight or Supplier, mere concentration of catalog Microplate, 384K, Greiner 784075 PS, white Protelna C-1 mg / ml, 115 kd (5 Calbiochem 236608 reactive, human, mere) recombinant from E. coli C1q, human 1 mg / ml, 410 kd Calbiochem 204876 Anti-CRP, rabbit 3.57 mg / ml, 165 kd Calbiochem 235762 Packard BioScience Detection Kit Alpha-Screen IgG 6760617 (Protein A) Packard BioScience Detection Kit Alpha-Screen IgG 6760607 Rabbit Anti-Cq1, Goat Calbiochem Polyclonal Serum 234390 Anti-streptavidin Sigma S6390 polyclonal serum Biotinila- Pierce 21430 Kit EZ-Link Sulfo-NHS-LC- Sodium Cireur 58.44 g / mol Merck 101540 Calcium chloride 147.02 g / mol Merck 102382 (CaCl2 x2 H20) Hydrochloride 157.6 g / mol Merck 108219 tris (hydroxymethyl) -aminomethane (Tris) Serum albumin Sigma P7888 bovine (BSA) Reagent / Plate Molecular weight or Supplier, number of concentration ro of catalog Phosphoryl chloride - 329.7 g / mol Sigma P0378 choline Gibco BRL saline solution buffered with 14200-067 phosphate (Mg + and Ca 2 * free) (PBS) Mini- Pierce P.69570 dialysis Slide-A-Lyzer Dimethylsulfoxide Merck KGaA, 1.02931 (DMSO) Ethanol Riedel de Haen, 32250 Plurónico F-68 Solution 10% Sigma, P5556 Hydrochloric acid 1 M Merck, 109057 Hydroxide solution 1 M Merck 109137 sodium HzO Milli-Q Preparation of the reaction buffer The following reaction buffer came in the Alpha-Screen system for use: 20 mM Tris-HCl, pH 7.2, 150 mM NaCl, 25 mM CaCl, 1 mM phosphorylcholine, 0.1 BSA %. For the preparation of a solution of Tris-NaCl buffer origin, 3.15 g of Tris-HCl and 8.77 g of NaCl in H20 were dissolved by 1 M NaOH, the pH was adjusted to 7.2 and then it was completed. with H2O up to 1 liter. The buffer was stored at room temperature. The reaction buffer was prepared by adding 36.7 mg of CaCb, 12.9 mg of phosphorylcholine and 50 mg of BSA to 50 ml of Tris-NaCl buffer.

BIOTINILATION OF ANTI-C1Q ANTIBODY In order to make possible an anti-C1q antibody binding to the streptavidin-coated donor granules, this antibody has to be biotinylated first. For this, 100 μ? of poly-clonal anti-C1q serum twice for one hour at room temperature against 200 ml of PBS. After adding 20 μ? from a 1.5 mM solution of sulfo-NHS (in milli-Q water) the solution was allowed to stand for 30 minutes at room temperature and then dialyzed again twice against PBS in order to separate the reagents in excess of biotinylation.

PERFORMANCE OF THE ASSAY To perform the test, 2 μ? of a donor granule / anti-CRP solution, then 2 μ? of C1q, 2 μ? of CRP and 2 μ? of acceptor / anti-CRP granules. The following final concentrations were adjusted: CRP: 1 nM; C1q: 10 nM; anti-CRP: 7.3 nM; Anti-C1q: 1: 1500; donor granules: 20 μg / ml; granules accept 40 pg / ml. As a negative control, reaction buffer was used in place of the CRP solution instead of the C1q solution or instead of these two solutions. After 2 hours of incubation at room temperature the results were read by means of an AlphaQuest reader.

RESULTS In the case of a CRP concentration of 1 nM and a C1q concentration of 10 nM, the following emitted radiation intensities (given in photomultiplier counts) were measured: In the absence of CRP and absence of C1q: 952 In the presence of CRP and absence of C1q: 1255 In the absence of CRP and presence of C1q: 1114 In the presence of CRP and presence of C1q: 80376 As can be recognized, negative controls do not provide a positive result, while by means of the joint addition of a solution containing C1q and CRP a strong signal can be recognized.

Claims

CLAIMS 1. - A method for determining a binding event of two components linked to CRP, directly or through one or more components, comprising the following steps: (a) initially introducing a solution containing CRP, (b) add to the solution containing CRP at least one donor component or a group of components, which contains at least one donor component, in the case of the donor component of a component comprising at least one compound or group of compounds, which they can emit a signal after excitation by a light source (donor group) and the donor component being able to bind to CRP, directly or through one or more components of the mentioned group of components, (c) add to the solution CRP containing at least one acceptor component or a group of components, which contains at least one acceptor component, in the case of the acceptor component of a component comprising and at least one compound or group of compounds, which capture the signal emitted by the compound or group of compounds according to (b) and can yield it in the form of electromagnetic radiation (acceptor group) and the acceptor component being in of binding to CRP, directly or through one or more components of the said group of components, (d) excite at least one compound or group of compounds according to (b) (donor group) by a light source, (e) detecting the electromagnetic radiation emitted by the at least one compound or group of compounds according to (c) (acceptor group) for the detection of the binding event. 2. A method for determining a binding event of two components bound to C1q, directly or through one or more components, comprising the following steps: (a) initially introducing a solution containing C1q, (b) add to the solution containing C1q at least one donor component or a group of components, containing at least one donor component, in the case of the donor component of a component comprising at least one compound or group of compounds, which they can emit a signal after excitation by a light source (donor group) and the donor component being able to bind C1q, directly or through one or more components of said group of components, (c) add to the solution containing C1q at least one acceptor component or a group of components, which contains at least one acceptor component, in the case of the acceptor component of a component that they buy of at least one compound or group of compounds, which capture the signal emitted by the compound or group of compounds according to (b) and can yield it in the form of electromagnetic radiation (acceptor group) and the acceptor component being in of binding to C1q, directly or through one or more components of said group of components, (d) excite at least one compound or group of compounds according to (b) (donor group) by a light source, (e) detecting the electromagnetic radiation emitted by the at least one compound or group of compounds according to (c) (acceptor group) for the detection of the binding event. 3. The method according to claim 1 or 2, characterized in that in the case of electromagnetic radiation it is a fluorescent radiation. 4. - The method according to claim 1 or 2, characterized in that in the case of the light source according to (d) it is a laser or a lamp. 5. The method according to claim 1 or 2, characterized in that the detection according to (e) is carried out with the help of a fo-tomultiplier. 6. - The method according to claim 1, characterized in that the components of the group of components according to (b) and / or (c) can be linked spatially successively to CRP or to each other. 7. - The method according to claim 2, characterized in that the components of the group of components according to (b) and / or (c) can be joined spatially successively to C1q or to each other. 8. - A method according to one of claims 1 to 7, characterized in that in the case of the components they are polypeptides, or they comprise a polypeptide. 9. - The method according to claim 8, characterized in that in the case of at least one of the components is an antibody. 10. The method according to claim 9, characterized in that in the case of the antibody it is a monoclonal or poly-clonal antibody. The method according to claim 1, characterized in that in the case of at least one of the components it is a participant in the natural union of CRP. 12. - The method according to claim 11, characterized in that in the case of the participant in the natural binding of CRP it is C1q. 13. - The method according to claim 12, characterized in that the group of components according to claim 1 (a) or claim 1 (b) comprises C1q and an anti-C1q antibody. 14. - The method according to claim 2, characterized in that in the case of at least one of the components is a participant in the natural union of C1q. 15. - The method according to claim 14, characterized in that in the case of the participant in the natural union is CRP. 16. The method according to claim 15, characterized in that the group of components according to claim 2 (a) or with claim 2 (b) comprises CRP and an anti-CRP antibody. 17. - The method according to claim 1 or 2, characterized in that the transmission of the signal by the at least one compound or group of compounds according to (b) (donor group) to the at least one compound or group of compounds according to (c) (acceptor group) takes place by energy transfer without emission. 18. - The method according to claim 7, characterized in that in the case of energy transfer without emission, it is a fluorescence resonance energy transfer. 19. - The method according to claim 1 or 2, characterized in that the signal transfer takes place by singlet oxygen. 20. - The method according to claim 19, characterized in that the at least one compound or group of compounds according to claim 1 (b) or 2 (b) (donor group) comprises a compound that, after excitation with a laser, it can transform the triplet oxygen into singlet oxygen, and the at least one compound or groups of compounds according to claim 1 (b) or 2 (b) (acceptor group) comprises at least one first compound, which can be excited by singlet oxygen, and comprises at least one second compound, which can pick up without energy the energy collected by the excitable compound by singlet oxygen and can emit it in the form of fluorescent radiation. 21. - The method according to claim 20, characterized in that the donor group and / or the acceptor group are located on particles. 22. - The method according to claim 21, characterized in that the particles have a mean diameter of about 200 nm. 23. - The method according to one of claims 1 to 22, characterized in that the method is used for the determination of the concentration of a solution containing CRP of unknown CRP content. 24. - The method according to claims 1 to 22, characterized in that the method is used for the determination of the concentration of a solution containing C1q of unknown C1q content. 25. - The method according to claim 23 or 24, characterized in that in the case of the solution is blood serum, blood plasma or blood serum or blood plasma diluted by a physiological buffer or with water. 26. - The method according to claim 25, characterized in that it is a diagnostic process for the detection of the state of inflammation of an organism and / or for the determination of the danger of heart infarction and / or apoplexy. 27. - The method according to one of the preceding claims, characterized in that it is a homogeneous method with HTS capability, which can be carried out by a single-stage process. 28. - The method according to claim 1, comprising, as an additional step, adding at least one test substance before step (a) and / or before step (b) and / or before the step (c) and / or before step (d), in order to observe whether this substance has an influence on the binding event between CRP and a component linked to CRP. 29. - The method according to claim 2, comprising, as an additional step, adding at least one test substance before step (a) and / or before step (b) and / or before the step (c) and / or before step (d), for the purpose of observing whether this substance has an influence on the binding event between C1q and a component attached to C1 q. 30. - The method according to claim 28 or 29, characterized in that the method is used for the purpose of determining substances that act by modulating, inhibiting or activating the binding event. 31.- A diagnostic test system with HTS capability, for the determination of a binding event between CRP and a component linked to CRP, which comprises the following components: (a) at least one donor component or a group of components that they contain at least one donor component, wherein the donor component of a component comprising at least one compound or group of compounds which, after excitation by a light source, can emit a signal (donor group), and the donor component able to join CRP, directly or through one or more components of the aforementioned group of components, (b) at least one acceptor component or a group of components, which contains at least one acceptor component, the component being acceptor of a component comprising at least one compound or group of compounds that captures the signal emitted by the compound or group of compounds according to (b), and can yield it in the form of electromagnetic radiation and, the acceptor component being able to bind to CRP, directly or through one or more components of the mentioned group of components. 32.- A diagnostic assay system with HTS capability for the determination of a binding event between C1q and a component linked to C1q, which comprises the following components: (a) at least one donor component or a group of components containing at least one donor component, wherein the donor component of a component comprising at least one compound or group of compounds which, after excitation by a light source, can emit a signal (donor group), and the component being donor able to bind C1q, directly or through one or more components of said group of components, (b) at least one acceptor component or a group of components, which contains at least one acceptor component, wherein the acceptor component is of a component comprising at least one compound or group of compounds that captures the signal emitted by the compound or group of compounds according to (b), and can yield it in the form of electromagnetic radiation and, the component being able to bind C1q, directly or through one or more components of the said group of components. 33. - The assay system with HTS capability according to claim 31 or 32 further comprising the following components: (c) blood serum or blood plasma or blood serum or blood plasma diluted in a physiological buffer or by water, (d) a light source for the excitation of at least one compound or group of compounds according to (a) (donor group), (e) a detection system for detecting electromagnetic radiation emitted in accordance with (b) . 34. - A diagnostic test system with HTS capacity for the determination of active substances, which act by modulating the interaction between CRP and a CRP-linked component, comprising the following components: (a) at least one donor component or a group of components containing at least one donor component, wherein the donor component of a component comprising at least one compound or group of compounds which, after excitation by a light source, can emit a signal (donor group), and the donor component being able to bind to CRP, directly or through one or more components of said group of components, (b) at least one acceptor component or a group of components, containing at least one acceptor component, wherein the acceptor component of a component comprising at least one compound or group of compounds that the signal emitted by the compound or group of compounds in accordance with (b) captures, and can transfer it in the form of electromagnetic radiation and, the component being and acceptor in conditions to bind to CRP, directly or through one or more components of the mentioned group of components, (c) at least one test compound. 35.- A diagnostic assay system with HTS capacity for the determination of active substances, which act by modulating the interaction between C1q and a component linked to C1q, which comprises the following components: (a) at least one donor component or one group of components containing at least one donor component, wherein the donor component of a component comprising at least one compound or group of compounds which, after excitation by a light source, can emit a signal (donor group) ), and the donor component being able to bind C1q, directly or through one or more components of the mentioned group of components, (b) at least one acceptor component or a group of components, which contains at least one component acceptor, whereby the acceptor component of a component comprising at least one "V" - 33 - compound or group of compounds that captures the signal emitted by the compound or group of compounds according to (b), and can yield it in the form of electromagnetic radiation and, the acceptor component being able to bind C1q, directly or through one or more components of the 5 mentioned group of components. (c) at least one test compound. 36.- Test system with HTS capacity according to claim 34 or 35, which also comprises the following components: D) a light source for the excitation of at least one compound or group of compounds according to (a) (donor group), a detection system for detecting the electromagnetic radiation emitted according to (b). fifteen twenty