DE19752712C2 - Method for measuring the association of substructures of pathological protein deposits - Google Patents

Description

The present invention relates to a method for Screening of active ingredients for the treatment of with patho logical protein-related diseases as well a procedure for the diagnostic detection of with patho logical protein-related diseases.

A number of different diseases are associated with the onset linked by pathological protein deposits. It is often unclear whether the protein deposits are only appearances of a clinical picture or whether these are protein deposits causally responsible for the disease itself as a pathogen are. So neurodegenerative diseases are known in such as those in the brains of those affected as amyloids Protein deposits designated as plaques are detectable. Such diseases include, for example Alzheimer's disease, bovine spongiform encephalopathy (BSE), Creutzfeldt-Jakob disease, Kuru disease, Scrapie as well as possibly other diseases that in the past also as "slow virus" diseases  were designated. Recently, the BSE Disease brought to the public consciousness, under other because BSE with Creutzfeldt-Jakob disease in People is associated. To date it is unclear what mechanisms the protein deposits into the Intervene in illness. The from is remarkable Prusiner observed association between infectiousness and the concentration of certain proteins in the disease happen scrapie, a neurodegenerative disease of sheep, play a role. Pathological protein Positioning does not only occur with diseases of the neuronal Systems in appearance, but also in others Organs observed, for example in a disease of the Type diabetes.

An overview of prion diseases is from D. Riesner in "Chemistry in our time" (1996), pp 66-74 published been. Among other things, it states that a safe and rapid diagnosis is a pressing problem for prions research represents, not just about biosafety to ensure, but also to basic research with many open questions about replication and pathogenesis to advance by prions. It is also desirable worth developing a screening process with which it succeeds in ver against drugs with protein deposits develop related pathological phenomena. Here should be both a screening process for detecting Active substances that deposit pathological proteins prevent, as well as to dissolve pathological protein deposits are provided.

Surprisingly, the task mentioned at the beginning solved by a method of screening with the features of claim 1. The diagnostic method according to the invention has the features of claim 2. Is common the two methods the principle of measuring the association  of substructures with the pathological protein storage.

Fig. 1 is a diagram showing the inventive method for the diagnosis of pathological protein depositions.

Fig. 2 shows the results of an experiment for the detection of prion-protein aggregates by means of fluorescence correlation spectroscopy.

Fig. 3 shows the results of an experiment for detecting prions isolated from tissue samples by fluorescence-labeled solubilized prion proteins (PrP-Cy2).

Fig. 4 shows the sensitivity of the homologous of fluorescently labeled solubilized prion proteins (PrP Cy2) of prion protein aggregates from tissue samples, and the effect of SDS concentration.

Fig. 5 shows the homologous detection of peptide aggregates, which represent the majority of the amyloid deposits in Alzheimer's disease, with fluorescently labeled soluble Aß (1-42) peptide.

Fig. 6 shows the results of an experiment for heterogeneous detection of peptide aggregates, which represent the majority of the amyloid deposits in Alzheimer's disease, with fluorescence-labeled solubilized prion protein (PrP-Cy2).

Fig. 7 shows the results of an experiment for speci fi c detection of aggregates in the cerebrospinal fluid of patients in whom Alzheimer's disease was diagnosed.

Fig. 8 is a diagram showing the inventive method for screening of active ingredients.

The screening method according to the invention for the detection of Active substances for the treatment of with pathological protein Deposits related diseases assume that in the presence of suspected active substances the association of Substructures of pathological protein deposits structures forming pathological protein deposits, structures corresponding to pathological protein deposits and / or pathological protein deposits as targets is measured. As an alternative, first an association from partial structures to pathological protein deposits forming structures, pathological protein deposits speaking structures and / or pathological protein Storage performed as targets and then the reverse the association under the influence of suspected active substances observed. Compounds are then identified as the active ingredient adorns who are able to reverse the association effect in a significant way. A third alternative is that the dissociation of protein deposits measured even under the influence of suspected active substances becomes.

The method according to the invention for diagnostic detection of diseases involves measuring the association of part structures of pathological protein deposits on patholo structures forming protein deposits, pathological structures and / or corresponding protein deposits pathological protein deposits.

According to the invention, the partial structures of the pathological Protein deposits, the pathological protein deposits forming structures, the pathological protein deposits corresponding structures and / or the pathological Protein deposits preferably have a detectable self shaft up.

The substructures are preferably the pathological ones Protein deposits monomeric or oligomeric units patho  logical protein deposits. The substructures can also homologous, in particular structural homologous, to monomers or oligomeric units of pathological protein deposits.

The substructures, their association with pathological Structures forming protein deposits, pathological Structures corresponding to protein deposits and / or patho Logical protein deposits can be measured from the same (homologous detection) or another type (hetero log detection) pathological protein deposits originate, which is used as a target. For example targeting pathological protein deposits Structure that corresponds to pathological protein deposits structure and / or pathological protein deposits itself that comes from a prion disorder like scrapie or BSE come from, whereas the substructures, whose association with the target is measured, for example can also come from structures other than those mentioned. So it is possible, for example, the association of part structures derived from amyloid plaques from BSE Protein deposits from tissues suffering from Alzheimer's measure up.

The substructures that are used according to the invention can preferably by treating pathological protein deposits, such as amyloid plaques, with physical Methods such as ultrasound treatment, exposure to tempera changes in the door, treatment with different solutions Ionic strength, treatment with solutions of chaotropic ions or Detergents and / or biochemical methods such as treatment the amyloid plaques with enzymes, especially proteases be preserved. So it is possible, for example, from on BSE diseased neuronal tissue due to amyloid plaques enzymatic degradation followed by ultrasound treatment in Presence of detergents to break down into substructures, which can subsequently build up fibrillar structures again. The partial structures of the pathological protein deposits  can also recombinant proteins, protein fragments or Be peptides which sequence homologies to the corresponding ones amyloid plaques of different origins and types can point.

As targets on which the association of substructures pathological protein deposits are measured structures into consideration. This includes structures understood, which itself is not the actual patholo represent protein deposits, but monomeric or oligomeric units of pathological protein deposits or larger aggregates of the substructures of the pathological Protein deposits, the association of which is to be measured, represent. Pathological protein deposits also occur appropriate structures into consideration. Such structures mimic Areas of the actual pathological protein deposits after that with the substructures according to the invention be placed, interactively interact. As The pathological proteins also come as an alternative storage itself.

The partial structures of the pathological protein deposits, the structures forming the pathological protein deposits, corresponding to the pathological protein deposits Structures and / or the pathological protein deposits preferably have a detectable property. The detectable property is either intrinsic in the Partial structure available or can be introduced later become. The at least one detectable property of Substructures are made especially by physical Methods, preferably recorded spectroscopically. As detect The property of the substructure comes from, for example, Size or dimension. Also features like Structure, measurable with circular dichroism, optical properties can such as luminescence, fluorescence or absorption for measuring the association of the substructures with the targets be used. For example, your own  fluorescence from substructures as well as from later marked substructures with fluorescent properties be used.

As methods for measuring the at least one detectable The characteristic of the substructure comes in particular from fluori metric methods such as confocal fluorescence spectroscopy, Fluorescence Correlation Spectroscopy (FCS), FCS in Com combination with cross correlation with corresponding ones Evaluation procedure into consideration. It is expressed here Lich on DE 44 38 341 and WO-A-96/13744, WO-A- 94/16313 and EP-A-96 116 373 and EP-A-97 109 353 referred. On the disclosure content of these registrations expressly referred. In particular, the application of the FCS cross correlation is advantageous because of this method the specificity of the method can be improved. In particular especially through the use of different substructures as probes or in combination with other probes for pathological protein deposits, such as. B. specific Antibodies can make false positive detection of aggregates be suppressed in biological media.

In cross-correlation, two are different fluorescent species - for example targets and part structures - observed.

The detectable properties are, for example, by Fluorescent labels, the low molecular weight, but also high molecular weight groups can be produced. So can for example, labeled antibodies attached to the targets be bound, mark the latter. Binding part structures can then be measured accordingly. It can different colored markings of different parts structures are made and a cross corrections on them lation be carried out. It is also possible that To provide substructures with appropriate labels that  either low molecular weight fluorescent ligands and / or are corresponding labeling conjugates.

According to the invention, the pathological protein deposits originate gene preferably from amyloid plaques that are associated with neuro degenerative diseases such as Alzheimer's disease, bovine spongiform encephalopathy (BSE), Creutzfeldt- Jakob disease, scrapie (scrapie), chorea Huntington's go along, or from amyloid plaques of others Organs as the neural system, such as with Diabetes-related protein deposits. A suitable one Provide source for pathological protein deposits Organ extracts, preferably brain extracts from diseased animals. For example, these can be Syrian infected with prions Be a golden hamster.

The partial structures of the pathological protein deposits can also recombinant proteins, protein fragments or Be peptides, which sequence homologies to amyloid plaques exhibit. For example, conservative amino acid Substitutions in highly conserved regions as follows are taken into account: each isoleucine, valine and leucine Amino acid can be made against another of these amino acids be exchanged, aspartate can be exchanged for glutamate and vice versa be exchanged, glutamine for asparagine and vice versa, Serine against threonine and vice versa. Conservative amino acid Substitutions in less highly conserved regions can be as follows: each of the amino acids isoleucine, valine and Leucine can be used against any other of these amino acids, aspartate against glutamate and vice versa, glutamine against asparagine and vice versa, serine against threonine and vice versa, glycine against Alanine and vice versa, alanine versus valine and vice versa, Methionine against any of the amino acids leucine, isoleucine or Valine, lysine against arginine and vice versa, one of the amino acids aspartate or glutamate against one of the amino acids Arginine or lysine, histidine against one of the amino acids Arginine or lysine, glutamine versus glutamate and vice versa  and asparagine for aspartate and vice versa his.

As a source of material that is a diagnostic sub search is subjected to the method according to the invention, body fluids such as liquor come in particular cerebrospinalis, blood, lymph, urine or secretions such as sputum in question. From the body fluids or secretions mentioned samples are taken and used to measure the association of potentially pathological contained in these samples Structures forming protein deposits, pathological Structures corresponding to protein deposits and / or pathological protein deposits, with substructures in Contacted and incubated. The presence of the pathological protein deposits that are indicative of a associated disease will result in a positive one diagnostic signal mediated by the association of the added substructures. Body fluids are against advantageous over tissues, because here a direct incubation can take place, whereas tissue is only opened up regularly must be closed. This can be done, for example mechanical or chemical treatment or combinations of which are done.

As parameters that result from the detectable properties of the Substructures are determined, especially trans lation diffusion speeds, rotational diffusion ge dizziness, lifetimes of excited states, pola radiation, energy transfer, quantum yield, Particle number or concentration, intensity differences in Consideration.

The following examples illustrate the process explained.

The diagram shown in FIG. 1 shows the experimental procedure for Examples 1 to 6 below.

The preparation of soluble (solubilized) prion Proteins (PrP) were sonicated in 10 mM Sodium phosphate pH 7.2, 0.2% SDS from prion aggregates, isolated from infectious brain tissue of Syrian gold hamsters (Riesner, D .; Kellings, K .; Post, K .; Wille, H .; Serban, H .; Groth, D .; Baldwin, M.A. and Prusiner, S.B., 1996, Journal of Virology, Vol. 70, No. 3, pages 1714-1722). Depending on the energy of the ultrasound system creates a monomeric to oligomeric PrP fraction, which for the following described examples (1 to 3, 5) fluores was zenzmarkiert (Pitschke, M .; Post, K. and Riesner, D, 1997, Progress in Colloid & Polymer Science, Vol. 107, in Print). The dye Cy2 (Amersham) used.

Soluble Aß (1-42) could in 10 mM Na phosphate buffer, pH 7.0 be stabilized with 0.2% SDS (Example 4, 6). For Fluorescent labeling of Aß also became the dye Cy2 (Amersham) used.

For association measurements, the soluble, monomeric to oligomeric fluorescence-labeled proteins or peptides with thinning of the SDS to 0.02% SDS with amyloids Aggregates (infectious prion particles: 1-3; Aß: 4-5) incubated and the fluorescence signal by means of fluorescence Correlation spectroscopy detected.

example 1 Detection of prion protein aggregates using fluorescence Correlation Spectroscopy (FCS)

Detection of infectious prion particles (PrP) using FCS by attachment of fluorescence-labeled solubilized PrP.  

Material:

• - Solubilized PrP-Cy2, approx. 25 ng / µl in 10 mM Na Phosphate buffer, pH 7.0, 0.2% SDS ⇒ (solution 1)
• - Prion particles (approx. 200 ng / µl) in 10 mM Na phosphate buffer, pH 7.0 ⇒ (solution 2)

Execution:

Two approaches were made:

• A) 1 µl solution 1 (1:10)
  19 µl Na phosphate buffer,
  pH 7.0
• B) 1 µl solution 1 (1:10)
  17 µl Na phosphate buffer,
  pH 7.0
  2 µl solution 2 (1: 100)

Both batches were mixed, incubated for 1 minute, in applied a sample chamber and measured in FCS.

Fig. 2 shows the result of the measurement of the fluorescence intensity (in relative units / RE) over a period of 60 seconds. In Fig. 2a to recognize the fluctuation signal of the solubilized PrP-Cy2 having a signal intensity 110-130 RE. An evaluation of the signal using autocorrelation results in a diffusion time of approx. 250 µs, which typically corresponds to the monomeric protein. FIG. 2b also shows a signal peak with a fluorescence intensity of approx. 950 RE, which is caused by the slow diffusion of a larger aggregate to which the previously solubilized fluorescent-labeled PrP-Cy2 is attached. Since it is only a one-off event, it is not possible to determine the diffusion time or the molecular weight using auto-correlation. However, a clear signal detection and assignment to individual aggregates succeeds via the appearance of isolated peaks with a drastic increase in the fluorescence intensity.

Example 2 Homologous detection of isolated prions from tissue samples with fluorescence-labeled solubilized prion proteins (PrP-Cy2)

Detection of infectious aggregates of prion proteins (PrP- Rods) in brain extracts after labeling with fluorescence labeled solubilized prion proteins (PrP-Cy2) using FCS.

Material:

• - Solubilized PrP-Cy2, approx. 25 ng / µl in 10 mM Na Phosphate buffer, pH 7.0, 0.2% SDS ⇒ (solution 1)
• - Brain extract from non-infected (solution 2) and prion infected Syrian golden hamsters (solution 3)

Execution:

Two approaches were made:

• A) 1 µl solution 1 (1:10)
  17 µl Na phosphate buffer,
  pH 7.0
  2 µl solution 2 (1:10)
• B) 1 µl solution 1 (1:10)
  17 µl Na phosphate buffer
  pH 7.0
  2 µl solution 3 (1:10)

Both batches were mixed, incubated for 1 minute, in applied a sample chamber and measured in FCS.

As a control, solubilized PrP-Cy2 without the addition of Measure brain extract in the same concentration.

The peak frequency for the individual batches is plotted in FIG. 3 as a graph. The number of peaks caused by diffusion of high molecular weight aggregates through the confocal volume element (see Example 1) was detected. It can be seen that the prion-infected tissue sample has a significantly higher peak frequency compared to the sample with non-infected brain extract.

Detection of infectious material by attachment of Solubilized PrP-Cy2 on existing prions is with this Experimental arrangement possible, even if the peak frequency at Examinations of non-infected tissue sample Shows underground.

Example 3 Sensitivity of homologous attachment of fluorescence labeled solubilized prion proteins (PrP-Cy2) and from Tissue samples isolated prion protein aggregates

The sensitivity of the homologous attachment of PrP-Cy2 on Brain tissue isolated infectious prions is analyzed with FCS siert.

Material:

• - Solubilized PrP-Cy2, approx. 25 ng / µl in 10 mM Na Phosphate buffer, pH 7.0, 0.2% SDS ⇒ (solution 1)
• - PrP rods (approx. 200 ng / µl) in 10 mM Na phosphate buffer pH 7.0 ⇒ (solution 2)

Execution:

Two series of tests were carried out:

• A) 1 µl solution 1 (1:10)
  17 µl Na phosphate buffer,
  pH 7.0,
  2 µl solution 2 (1: x; amount of PrP, see Fig. 4)
• B) 1 µl solution 1 (1:10)
  17 µl Na phosphate buffer,
  pH 7.0, 0.2% SDS
  2 µl solution 2 (1: x, (PrP amount see Fig. 4)

The amount of PrP-Rods in the test approach was stepped up lowered to the detection limit. The measurement in 0.2% SDS serves as a control.

All batches were mixed, incubated for 1 minute, in a Sample chamber applied and measured using FCS.  

The peak frequency, expressed in peaks per 10 min measuring time, was plotted in FIG. 4 as a function of the amount of PrP in the test batch. A direct linear dependence on the peak frequency can be seen over the measured concentration range from 40 ng to 400 pg. The evaluation of the number of peaks allows a direct quantification of the prion concentration in the sample to be measured. In 0.2% SDS, installation in existing PrP structures is not possible.

Example 4 Homologous detection of peptide aggregates that do the bulk the amyloid deposits in Alzheimer's disease represent with fluorescently labeled soluble Aß (1-42) - peptide

Detection of aggregates of Alzheimer's disease with fluorescent-labeled soluble Aß (1-42) using FCS with the Question whether the one used in Examples 1 to 3 Detection method also for other diseases with patho logical protein deposits is transferable and thus to Detection of any amyloid structure is suitable.

Material:

• Soluble Aß peptide 1-42, fluorescently labeled with Cy2, 100 ng / µl in 10 mM Na phosphate buffer, pH 7.0, 0.2% SDS ⇒ (solution 1)
• - Aß amyloid fibrils (90 ng / µl) in 10 mM Na phosphate buffer pH 7.0 ⇒ (solution 2)

Execution:

Two approaches were made:

• A) 1 µl solution 1 (1:10)
  10 µl solution 2
  9 µl Na phosphate buffer,
  pH 7.0
• B) 1 µl solution 1 (1:10)
  10 µl solution 2
  9 µl Na phosphate buffer,
  0.2% SDS, pH 7.0

Both batches were mixed, incubated for 1 min, in one Sample chamber applied and measured in the FCS.

As a control, soluble Aß (1-42) -Cy2 was used in the same Measure concentration.

The results of the experiment are summarized graphically in FIG . The evaluation of the peak frequency (cf. 1-3) shows that when incubated in 0.01% SDS, Aß fibrils are labeled (identical to PrP aggregates), which are the main components of the amyloid deposits in Alzheimer's disease this procedure is possible. Incorporation of Aß-Cy2 in the presence of an increased SDS concentration (0.1%) is not possible. It is also not possible to label Aß which is in a soluble (non-fibrillary) structure. The experiment shows that detection of amyloid deposits associated with Alzheimer's disease is possible.

Example 5 Heterologous detection of peptide aggregates that do the bulk the amyloid deposits in Alzheimer's disease represent with fluorescence-labeled solubilized prion Proteins (PrP-Cy2)

Detection of aggregates of Alzheimer's disease with fluorescent-labeled soluble Aß (1-42) using FCS with the Question whether a heterologous label with a fluorescent-labeled soluble sample is not possible is identical to the sample to which an attachment takes place should find.  

Material:

• - Solubilized PrP-Cy2, approx. 25 ng / µl in 10 mM Na Phosphate buffer pH 7.0, 0.2% SDS ⇒ (solution 1)
• - Aß (1-42) amyloid fibrils (90 ng / µl) in 10 mM Na Phosphate buffer pH 7.0 ⇒ (solution 2)
• - Ate (1-42) non-fibrillary (90 ng / µl) in 10 mM Na Phosphate buffer pH 7.0, 0.2% SDS ⇒ (solution 3)
• - Aß (1-40) amyloid fibrils (90 ng / µl) in 10 mM Na Phosphate pH 7.0 ⇒ (solution 4)
• - Ate (1-40) non-fibrillary (90 ng / µl) in 10 mM Na Phosphate buffer pH 7.0, 0.2% SDS ⇒ (solution 5)

Execution:

The following approaches were made:

Aß peptide (1-42):

• A) 1 µl solution 1 (1:10)
  2.5 µl solution 2
  16.5 µl Na phosphate buffer,
  pH 7.0
• B) 1 µl solution 1 (1:10)
  2.5 µl solution 3
  16.5 µl Na phosphate buffer,
  pH 7.0
• C) 1 µl solution 1 (1:10)
  2.5 µl solution 2
  16.5 µl Na phosphate buffer,
  pH 7.0, 0.2% SDS

Aß peptide (1-40):

• A) 1 µl solution 1 (1:10)
  2.5 µl solution 4
  16.5 µl Na phosphate buffer,
  pH 7.0
• B) 1 µl solution 1 (1:10)
  2.5 µl solution 5
  16.5 µl Na phosphate buffer,
  pH 7.0
• C) 1 µl solution 1 (1:10)
  2.5 µl solution 4
  16.5 µl Na phosphate buffer,
  pH 7.0, 0.2% SDS

All batches were mixed, incubated for 1 min, in a Sample chamber applied and measured in the FCS.

As a control, soluble PrP-Cy2 was used in the same con measure the concentration at 0.01% SDS.

The peak frequency (see Example 2) was measured for each of the approaches listed above ( FIG. 6). It shows that both Aß (1-42) and Aß (1-40) can be marked clearly with PrP-Cy2 if they are in a fibrillary structure, and the marking reaction takes place at 0.01% SDS. When attached to non-fibrillar structures and with an increased SDS concentration, only a small background signal can be seen.

This result shows that it is for amyloid labeling Protein aggregates is not absolutely necessary for that Label used soluble fluorescent labeled protein must be identical to the aggregate-forming protein.

Example 6 Detection of aggregates in the cerebrospinal fluid of patients participating in the Suffer from Alzheimer's disease

Specific detection of aggregates using the Alzheimer's disease. This is liquor (Spinal fluid) used as a sample in which spec fish between patients suffering from Alzheimer's Er disease and a healthy control group can be divorced.

Material:

• Soluble Aß peptide 1-42, fluorescently labeled with Cy2, 100 ng / µl in 10 mM Na phosphate buffer pH 7.0, 0.2% SDS ⇒ (solution 1)
• - CSF from diseased patients (2x) and control patient not ill (1x)

Execution:

The following approaches were made:

• A) 2 µl solution 1 (1:10)
  18 µl of CSF from patient 1 (healthy)
• B) 2 µl solution 1 (1:10)
  18 µl CSF from patient 2 (diseased)
• C) 2 µl solution 1 (1:10)
  18 µl liquor from patient 3 (diseased)

All batches were mixed, incubated for 1 min, in a Sample chamber applied and measured in the FCS.

The peak frequency (see Example 2) was measured for each of the approaches listed above ( FIG. 7). It shows that aggregates were clearly detected in the cerebrospinal fluid of patients suffering from Alzheimer's disease. No background signal could be detected in the CSF of the healthy patient. The experiment shows that there is a clear correlation of the measurement signal with Alzheimer's disease.

Claims (14)

1. A method for screening active substances for the treatment of diseases which are associated with protein deposits, by measuring the association of substructures of the pathological protein deposits with structures forming pathological protein deposits, structures corresponding to pathological protein deposits and / or pathological protein deposits as targets in the presence of suspected active substances, or
Measurement of the association of substructures of pathological protein deposits with structures forming pathological protein deposits, structures corresponding to pathological protein deposits and / or pathological protein deposits as targets and measurement of the reversal of the association in the presence of suspected active substances or
Measurement of a dissociation of pathological protein deposits, pathological protein deposits forming structures and / or pathological protein deposits of corresponding structures in the presence of suspected active substances. 2. Procedure for the diagnostic detection of Er diseases associated with protein deposits, by measuring the Association of substructures of the pathological Protein deposits, pathological protein deposits forming structures, pathological protein deposits corresponding structures and / or pathological Protein deposits, on partial structures of the pathological protein deposits, structure forming pathological protein deposits corresponding pathological protein deposits  Structures and / or pathological protein deposits as targets. 3. The method according to any one of claims 1 and / or 2, wherein the substructures of the pathological protein deposits, the pathological protein deposits forming structures, the pathological protein deposition structures and / or the patho logical protein deposits at least one have detectable property. 4. The method according to any one of claims 1 to 3, wherein the Substructures of pathological protein deposits with at least one detectable property mono mer or oligomeric units of pathological protein are deposits or the substructures are homologous pathological to monomeric or oligomeric units Protein deposits. 5. The method according to any one of claims 1 to 4, wherein the Substructures, their association with pathological Pro structures that build up the deposits, pathological Structures corresponding to protein deposits and / or pathological protein deposits are measured from the same or a different type of pathological Protein deposits originate, which ver as target is applied. 6. The method according to any one of claims 1 to 5, wherein the Substructures through treatment of pathological protein deposits, such as amyloid plaques, with physical Methods such as ultrasound treatment, exposure to Temperature changes, treatment with solutions below different ionic strength, treatment with solutions chaotropic ions and / or chemical methods such as Treatment of amyloid plaques with enzymes, esp special proteases, or detergents are available.   7. The method according to any one of claims 1 to 6, wherein the at least one detectable property either be is already present in the substructure or later is introduced. 8. The method according to any one of claims 3 to 7, wherein the at least one detectable property size or Expansion, molecular weight, structure, circular dichroism, optical property such as luminescence, in particular Fluorescence, absorption is. 9. The method according to any one of claims 3 to 8, wherein the Measurement of the at least one detectable property using physical methods, in particular spectroscopic methods. 10. The method according to any one of claims 3 to 9, wherein the Measurement of the at least one detectable property using fluorimetric methods such as confocal Fluorescence spectroscopy, fluorescence correlation Spectroscopy (FCS), FCS in combination with cross correlation with the respective evaluation procedure takes place. 11. The method according to any one of claims 1 to 10, wherein the pathological protein deposits amyloid plaques, those with neurodegenerative diseases, such as Creutz Feldt-Jakob disease, bovine spongioform Encephalopathy (BSE), scrapie (scraping), Huntington's disease, Alzheimer's disease go, are, or from amyloid plaques of other organs than the neural system, like that associated with diabetes Protein deposits. 12. The method according to any one of claims 1 to 11, wherein the Substructures of pathological protein deposits recombinant proteins, protein fragments or peptides  are the sequence homologies to amyloid plaques point. 13. The method according to any one of claims 2 to 12, wherein from Body fluids such as cerebrospinal fluid, blood, Lymph, urine, or secretions such as sputum or tissues Samples taken and for diagnostic purposes any diseases with substructures of the patho logical protein deposits, pathological protein deposits forming structures, pathological Structures corresponding to protein deposits and / or brought into contact with pathological protein deposits, incubated and their association measured on targets becomes. 14. The method according to any one of claims 2 to 13, wherein from the detectable properties parameters such as trans diffusion velocities, rotation diffusion speeds, lifetimes excited States, polarization of radiation, energy transfer, Quantum yield, particle number or concentration, Differences in intensity are determined.