DE102009035502A1 - Method and device for detecting the movement and attachment of cells and particles to cell, tissue and implant layers in the simulation of flow conditions - Google Patents

Abstract

The task was to record the movement and deposition processes of cells and particles more precisely and completely when simulating flow conditions. According to the invention, the movement and accumulation of cells and particles in the at least one interaction zone (2, 3, 4) are completely optically monitored and evaluated, transmigrating cells and particles are collected for control and evaluation (7, 8, 9) and, in particular, the culture conditions for long-term studies of the flow medium for flowing through the interaction zone (2, 3, 4), for example by adjusting the pH value, the temperature and the nutrient supply for the cells and particles. The invention is used, for example, for testing pharmaceuticals, pathogens and other active substances.

Description

The The invention relates to a method and a device for detecting motion and attachment processes of cells and particles to cell, Tissue and implant layers in the simulation of flow conditions capture. Such movement and attachment processes The cells and particles, especially of blood cells, are significant for the study of cell and tissue physiology and of Effects caused by drugs, pathogens or other active substances be evoked and the behavior of these cells and particles affect their flow through the organism.

to Determination of major and side effects of these drugs, various Pathogens and other agents on the said organism is therefore their influence on the accumulation, migration and Differentiation behavior, including other medical or of biologically relevant behaviors and reactions, of suspension and adherent cells on cell and tissue surfaces under flow conditions in simulation devices outside of the organism.

The simulation devices should be able to reproduce the flow conditions occurring in the body as realistically as possible in order to be able to make organism and tissue-relevant statements. With these simulation methods, in particular animal experiments are to be substituted or at least limited, which allow life-like analysis in the complex system of the organism taking into account many physiologically relevant parameters, but ethically controversial, often find no acceptance and thus more and more come under criticism. Moreover, in animal models, real-time observation of disease development and disease progression is limited. In addition, it is often necessary to kill the treated animals at the end of the experiment. As a result, only a snapshot of the course of the disease is made possible. Due to the individual differences that occur in inbred animals, it is almost impossible to reconstruct a representative disease course even in a large number of examinations and killed experimental animals. Furthermore, the transferability of the findings obtained in the animal model to humans is usually questionable ( TM Bocan: Animal models of atherosclerosis and interpretation of drug intervention studies, Curr Pharm Des 4, 1998, 37-52 ; AC McMahon, L. Critharides and HC Lowe: Animal Models of Atherosclerosis Progression: Current Concepts, Curr Drug Targets Cardiovasc Haematol Disord 5, 2005, 433-440 ; JC Russell and S. Proctor: Small animal models of cardiovascular disease: tools for the study of the roles of metabolic syndrome, dyslipidemia, and atherosclerosis. Cardiovascular Pathol 15, 2006, 318-330 ).

To simulate the flow conditions in the organism, a flow model for the investigation of the attachment processes of cells is known ( DE 100 19 833 C2 ), in which a tail pipe at an angle greater than 0 ° points to an interaction surface. Through this experimental arrangement, turbulent flows are created in this flow model. By means of an observation unit, consisting of a mounted parallel to the tailpipe microscope with condenser, the cell attachment processes are observed in the flow medium to the interaction surface.

The Microscope with condenser is on a focal plane above the interaction surface directed and can only look at this level. The observation but is limited to this defined level, so that the cells in their movement are not continuous in real time can be tracked. In the considered plane it is only possible, the cells essentially as static Size of single measurements to capture.

The in the DE 100 19 833 C2 deliberately selected flow rate in the flow chamber can be due to the device, the setting of flow and pressure conditions only very limited, in particular can be built and evaluate laminar flow conditions at the interaction surface. Single observations of cells are not spatially resolvable due to the high dynamics of the cell movement.

Furthermore This device forms only the flow conditions the bloodstream and can thus observe cells that are in Move this simulated bloodstream and hang on the wall if necessary stay and attach. Cells on the other hand, which walk through this wall and thus can be found below the interaction area, can not be recorded and evaluated and are thus the Investigation inaccessible. Such a statement is but for the analysis of, for example, a drug delivery for Pharmaceuticals of great interest.

In addition, especially for long-term studies, important environmental parameters, such as sterility, pH, etc., to comply with the physiology of the organism to be reproduced, can not be maintained. Morphological adaptations of cells to the applied flow conditions take several days. Thus, a consideration of the direct effects of flow on the morphology and physiology of the cells is not given. Furthermore, the device offers no possibility to investigate multilayer cell aggregates. Furthermore, the device does not allow the multiple Examine particles and cells.

It is also known to detect adhesion and migration processes of T cells under flow conditions in a system. (eg G. Cinamon and R. Alon: A real time in vitro assay for studying leukocyte transendothelial migration under physiological flow conditions, J. Immunol. Methods 273 (1-2), 2003, 53-62 ; T. Schinder, V. Shinder et al .: Shear flow-dependent integration of apical and subendothelial chemokines in T-cell transmigration: implications for locomotion and the multistep paradigm, Blood 109 (4), 2007, 1381-1386 ).

In Both devices become single cell layers according to the same principle from endothelial cells in commercial cell culture inserts cultured. Before the start of the experiment, the endothelial cells with the incubated substances to be tested by placing these directly on the cells are given. Subsequently, the substances are washed away and the actual attempt started. The flow of the medium is over produces a syringe pump. Adhesion and transmigration be by a perpendicular angle above the interaction surface attached phase-contrast microscope observed.

Endothelial cells need time to adapt to flow conditions. Under static conditions grown endothelial cells have a different physiology than cells that arise under the influence of flow conditions (eg. JT Butcher, AM Penrod, AJ Garcia and RM Nerem: Unique morphology and focal adhesion development of valvular endothelial cells in static and fluid flow environments, Arterioscler Thromb Vasc Biol 24, 2004, 1429-1434 ).

The Devices do not provide a requirement for cells under flow conditions to attract. There is no possibility for longer cultivation given by endothelial cells under flow conditions and the results are very limited compared with the in vivo situation.

The transmigrated cells are collected in a container. However, when penetrating the interaction surface remain Transmigrated cell adhere to the underside, which is disregarded in the evaluation stay. Gravity is not enough for all to be transmigrated Cells fall down to complete quantitative evaluation to be able to. These are adhesion and migration processes only inaccurately detectable.

Furthermore none of the devices allows repeated examination the cells and particles in their flow through the interaction zone. Also Cells and particles in the interaction zone can only on the apical side before the start of the experiment with the substances to be tested be incubated.

Also known is a system for monitoring adhesion and migration processes by means of a capillary system. Tumor cells are inserted into this capillary system and passed over a monocell layer. Adhesion processes are also monitored by a microscope on the interaction surface. For this purpose, a commercial 48-well cell culture plate is used ( C. Dong, MJ Slattery et al .: In vitro characterization and micromechanics of tumor cell chemotactic protrusion, locomotion, and extravasation, Ann Biomed Eng 30 (3), 2002, 344-55 ).

Also Here, transmigrating cells can become adhesions not be recorded and analyzed. In addition, the cell examination given only on said monocell layer.

The Pressure and flow conditions in the capillary system are influenced only limited. Also with this method are no long-term studies possible.

In further publications ( JJ Chiu, DL Wang et al .: Effects of disturbed flow on endothelial cells, J Biomech Eng 120 (1), 1998, 2-8 and JJ Chiu, CN Chen et al .: Analysis of the effect of disturbed flow on monocytic adhesion to endothelial cells, J Biomech 36 (12), 2003, 1883-95 ), the authors describe a system similar to that of Cinamon and Alon (2003) as well as from Schreiber and Shinder et al. (2007) mentioned. The system is supplemented by the possibility of detecting fluorescence. It consists of an inverted fluorescence microscope, which points to a flow chamber consisting of a channel. To the flow chamber, a pump system is connected. In addition to the disadvantages already mentioned above, there are no possibilities for perfusion below the interaction surface.

Also here is in particular a longer cultivation of cells excluded under river conditions, opportunities for Adjustment and adjustment for temperature and pH exist Not. Transmigrating cells can not be detected and are not accessible for a later evaluation.

In the patent family too CA 2 503 203 A1 describes a chamber system with a plurality of chambers, wherein at least the first chamber is connected by a channel with a second chamber. Through this channel, a liquid flows through a cell layer of endothelial cells. There is an observation unit for the detection of cell adhesion and cell migration.

As already described in the aforementioned systems (cf. Cinamon and Alon, 2003, and Schreiber and Shinder et al., 2007 ), even in this device transmigrating cells evade after adhesion of the observation and the subsequent evaluation. Again, only monolayers are used for investigation. Real-time analysis of cell observation is also not possible. The adjustment and adjustment of temperature and pH are not given. The cell layer can be incubated with the substances to be tested only from the apical side before starting the experiment. No dynamic perfusion is possible during the experiment.

In summary It should be noted that the known methods of simulation and Analysis of movement and attachment processes of the cells and particles on cell and tissue layers that are the living organism come close, are inadequate. Adhesion and Migration processes of cells under flow conditions are, if at all possible, only very limited and with faulty quantitative assessment possible. Long-term studies under the aforementioned conditions, in particular for Analyzes of pharmaceuticals and other active substances are important such an evaluation largely closed or show little Relevance to the transferability of the results to the living organism, so that despite all simulation methods animal experiments in spite of all ethical problems still an important meaning, especially for biology, medicine and drug research exhibit. The disadvantages of animal experiments are in particular in that the animals during or at the end of the experiments be killed and that a reproducible local effect of pathogens, drugs and other agents on disease progression at defined points of the body even when using of inbred strains, due to individual differences the individual animals used, very difficult to reproduce are. This leads to the necessity of implementation from a variety of individual experiments to address this disadvantage To balance the use of statistical tools. But even that Single observation of flowing and accumulating cells in standardized examination devices, as they currently are is known prepares due to the determined on a plane visual observability and the high dynamics of cell movement biggest trouble, so the evaluation essentially to the collection of statistical quantities amounts.

Of the The invention is therefore based on the object, the movement and Anlagerungsvorgänge of Cells and particles more accurate in the simulation of flow conditions and completely grasp.

On the one hand should the movement and attachment processes under conditions can be simulated and evaluated, which correspond to the pressure and flow behavior in the organism or body's own vessel also for Long term investigations come much closer. on the other hand should also be the adhesion and migration behavior the cells and particles as completely as possible and can be detected with high evaluation accuracy.

With The invention relates to the movement and attachment of the cells and particles, including their migration through the cell, tissue and implant layer, throughout the at least one interaction zone (and not only by looking from above) optically recorded and evaluated. The detection is preferably carried out by a plurality of observation beam paths, for example, by two stereomicroscopy systems, spatially visually, so that the interaction zone in an area not only above, but also inside and possibly below it visually spatially resolved and completely monitored. With this monitoring can also simultaneously a spectroscopic Detecting the movement and attachment of cells and particles, including the Interaction zone with its cell, tissue and implant layers itself, connected to at least one of said observation beam paths a spectroscopy unit is coupled.

By the cell, tissue and implant layer transmigrating this at least one interaction zone Cells and particles which are optically traced in this way and may optionally be analyzed spectrometrically be in at least one perfusion chamber below the interaction zone collected and are thus a further evaluation fed. Especially for long-term studies, the flow conditions the flow medium for the passage of the interaction zone, For example, by adjusting the pH, the temperature, oxygen saturation and nutrient supply for the cells and particles, specially adjusted, for example by a metered control.

With the invention described herein, the possibility is created, movement, adhesion and Transmigrationsprozesse of cells under flow conditions, as z. B. present in blood and lymph vessels of a living organism, to replicate almost lifelike, these processes spatially and temporally dissolve and isolate desired target cells vital after the study and thus to make these cells / particles for subsequent studies available. Furthermore, cells / particles under near-natural conditions can be analyzed spectrometrically without contact and label. The invention integrates components of the cell and tissue culture and optical imaging and processing and is therefore versatile.

investigations to cells of the blood and blood vessels often performed under static conditions. However, this does not correspond to the situation in a living organism, in which these cells are exposed to different flow conditions. The cells adapt to the flow and show under flowing Conditions deviating cell physiological properties against static Conditions. Investigation of cells adapted to the flow give thus more exact information about the behavior and the Characteristics of cells in and on the blood vessel in vivo. The invention allows investigations of this kind among various Types of flow conditions (turbulent, laminar, pulsating) and also allows the regulation of important physiological Parameters such as hydrostatic pressure, flow velocities, cell / particle concentration in the flow medium, pH and oxygen saturation. It is possible, whole tissue layers either in cell culture vorzuzüchten and integrate into the invention or the Growing of the tissue already under flow conditions in the system perform. For this purpose, the invention allows an individual Adaptation of physiological or pathological flow conditions for the cells to be examined over desired Periods. So this system allows for the one, the free controllable addition of various substances not only from the apical but also from the basolateral side of the cell / tissue or Implant layer. This is important because z. B. subcellular bioactive substances such as modified LDL, a strong influence on the interaction of leukocytes with the have entire tissue association. In addition, with this invention, the investigation the effects of various drugs, biologically active molecules, chemical pollutants and microbiological organisms, such as bacteria, parasites and fungi, not just the interaction of cells in the flow medium with the apical cells of the vessel wall limited, but also allows an investigation of the vascular wall transmigrating and / or in the tissue differentiating cells. By means of integrated in the invention Perfusion chambers and the cell traps they contain, it is possible completely migrated cells to catch vital and subsequent accessible to analytical and / or functional studies close. Furthermore, the invention has over a freely controllable control of the environmental parameters pH value, temperature, Amount of oxygen and nutrient addition, thus physiological Conditions can be set or by the adjustable Environmental parameters related pathological events, such as alkalosis, Acidosis, nutrient restriction, hypoxia standardized and can be analyzed close to the in vivo situation.

Diseases, like cancer and sepsis, whose spread occurs through blood flow, can with this invention much more accurate, more thorough and more extensively than with previous in vitro methods become. The invention makes it possible to metastasize of tumor cells cost-efficient, standardized and without use of animal models. Here, physiological Conditions are simulated to give a comprehensive picture of to maintain the spread of tumor cells in vivo. It is possible through the use of different tissue associations, different Imitate tissue areas of the body and thus metastasis of certain tumors in defined tissues. moreover It is possible to study for a long time To carry out periods and with one or more times Medium circulation to work within the system. This will be the opportunity created long-term pathophysiological processes, such as the emergence of atherosclerotic lesions, in real time. Furthermore, it is possible the effect of pharmacologically active substances, particles or cells in the To document the course of pathophysiological processes in real time. This may allow studies on the development of interventional therapies be made more effective, since drug concentrations are dynamic can be changed and the impact on pathophysiological processes can be observed directly. These possibilities open up, for example the study of atherosclerotic lesions, with it associated inflammatory reactions and pharmacological action effective substances provide new opportunities since cells are human Origin can be used and on the use can be dispensed with animal models. This allows processes relevant to the pathogenesis in humans be better examined. The advantages of the invention come in a similar way also in the study of other disease-relevant Events (eg sepsis, cancer, autoimmune diseases etc.) to wear on and in the tissue. Among the advantages of the invention continue the possibility of adhesion and transmigration of cells both in the eye and on the side and in the tissue capture. To achieve this, stereomicroscopes are used, which are also able to detect fluorescence signals. By the use of fluorescent dyes in the long-wave infrared range emit, it is possible cell movements also within of tissues. By introducing horizontal transparent Intermediate layers in tissues to be examined, it is still possible Also to detect cell movements in the gray field by means of phase contrast.

The invention will be described below with reference to a chamber body shown in the drawing as an exemplary embodiment in different Ansich be explained in more detail.

It demonstrate:

1 : Chamber body with three arranged below the interaction zone perfusion chambers in a sectional schematic side view

2 : frontal view of the chamber body with indicated objective lens and tube system

3 : lateral view of the chamber body in the longitudinal axis with indicated supervisory and lateral lens and tube systems including fluorescence unit

4 : Side view of the chamber body in the longitudinal axis with indicated objective lens and tube systems and lateral Raman microscope and laser

5 : schematic representation of the liquid circulations

1 shows a chamber body 1 with three interaction zones 2 . 3 . 4 , each consisting of a replicated to the human body cell, tissue and implant layer. The chamber body 1 has an inflow 5 on, over which a flow medium 6 with the cells and particles to be examined (indicated by the arrow) in the chamber body 1 is introduced so that the cells and particles of the incoming flow medium 6 successively the interaction zones 2 . 3 . 4 , under which three perfusion chambers 7 . 8th . 9 are arranged, flow through and come into contact with the respective cell, tissue and implant layer. After flowing through the chamber body 1 with the interaction zones 2 . 3 . 4 the flow medium enters via a drain 10 out again. The chamber body is via an integrated heating / cooling water duct system 11 tempered.

The interaction of the cells and particles of the flow medium with the interaction zones 2 . 3 . 4 can by a laterally or prudently arranged microscope system 12 be observed (cf. 2 ). As a microscope system 12 Both stereomicroscope systems and confocal laser scanning microscopes or other microscopy and spectroscopy systems can be used in a manner known per se. The observation of the interaction zones 2 . 3 . 4 through the microscope system 12 done by in the chamber body 1 and the interaction zones 2 . 3 . 4 associated replaceable glass or quartz inserts 13 . 14 . 15 , For complete observation of the totality of all interaction zones 2 . 3 . 4 is the microscope system 12 about (for reasons of clarity not shown in the drawing means, for example, known rail-like guide elements for transversal position change and also known per se pivot or rotation elements for changing the angular position, on the side of the chamber body 1 with the glass or quartz inserts 13 . 14 . 15 traversable.

Through the microscope system 12 may be the adhesion, migration and differentiation of the cells and particles of the flow medium as well as the cell, tissue and implant layers of the interaction zones 2 . 3 . 4 be observed and evaluated in real time.

In 3 are two microscope systems 12 representing the interaction zones 2 . 3 . 4 of the chamber body 1 both overseer and laterally (observation beam path 36 ) observe. The two microscope systems shown separately for reasons of clarity 12 For example, by separate and preferably in a right angle to each other arranged microscope systems or by a single microscope system with corresponding observation beam paths (with supervisory beam path 37 and lateral beam path 36 to the chamber body 1 attacking).

For the evaluation and documentation of the microscope systems shown separately 12 detected signals are the two microscope systems 12 each via a camera port 16 respectively. 17 with a computer system 18 coupled (cf. 3 ).

For the detection of fluorescence signals on cells and particles of the flow medium and / or on the said cell, tissue and implant layers of the interaction zones 2 . 3 . 4 and / or the perfusion chambers 7 . 8th . 9 these areas are illuminated by a suitable and fluorescent dyes stimulating light source 19 irradiated.

For the detection of metabolic processes on cells and particles of the flow medium and / or on the said cell, tissue and implant layers of the interaction zones 2 . 3 . 4 and / or the perfusion chambers 7 . 8th . 9 In addition, spectroscopy measurements can be carried out. For this purpose, the optical excitation is carried out by a laser 20 (see. 4 ).

The measurement of the spectra is carried out by a detector 21 , which can be integrated in the system, for example in the form of a known Raman microscope, the detector 21 is for signal digitization to a transducer system 22 coupled with the computer system 18 for data analysis.

For introducing cells or particles into the flow medium, a cell reservoir is used 23 connected to a heatable / coolable medium reservoir 24 is coupled (see schematic illustration 5 ). Control of pH value and pO ₂ value can be effected by gassing plants, which are in the medium reservoir 24 or in the cell reservoir 23 can be integrated (for reasons of clarity not explicitly shown in the drawing). An addition of substances into the flow medium may be through a substance application valve 25 respectively. The cell reservoir 23 , the medium reservoir 24 and the substance application valve 25 are in a medium cycle 26 involved. 5 shows this medium cycle 26 , in which also a sensor chamber 27 with sensor elements for the detection of cell culture parameters, such as pH, temperature and nutrient supply, for those in the flow medium 5 existing cells and particles is arranged. In this way, the said cell culture parameters in the medium cycle 26 controlled and can, for example, as mentioned above, via the mentioned gassing of the medium reservoir 24 or the cell reservoir 23 be controlled application-specific.

The temperature of the chamber body 1 via a heating / cooling circuit 28 , At which a temperatureable Heizwasserreservoir 29 connected to the heating / cooling circuit 28 fed.

Below the chamber body 1 there are three perfusion chambers (corresponding to the three said interaction zones) on the same 30 . 31 . 32 arranged (see also perfusion chambers 7 . 8th . 9 in 1 ), to each of which a reservoir 33 . 34 . 35 for additional in the medium circulation 26 introduced test substances and other applications is connected.

LIST OF REFERENCE NUMBERS

1

chamber body

2, 3, 4

Interaction zone

5

inflow

6

inflowing flow medium

7, 8, 9

perfusion chamber

10

outflow

11

Heating / cooling water channel system

12

microscope system

13 14, 15

Glass- or quartz insert in the chamber body 1

16 17

camera port

18

computer system

19

light source

20

laser

21

detector

22

converter system

23

cell reservoir

24

medium reservoir

25

Drug application valve

26

Medium circuit

27

sensor chamber

28

Heating / cooling cycle

29

Heizwasserreservoir

30 31, 32

perfusion chamber

33 34, 35

reservoir

36 37

Observation beam path

QUOTES INCLUDE IN THE DESCRIPTION

This list The documents listed by the applicant have been automated generated and is solely for better information recorded by the reader. The list is not part of the German Patent or utility model application. The DPMA takes over no liability for any errors or omissions.

Cited patent literature

• - DE 10019833 C2 [0004, 0006]
• - CA 2503203 A1 [0020]

Cited non-patent literature

• TM Bocan: Animal Models of Atherosclerosis and Interpretation of Drug Intervention Studies, Curr Pharm Des. 4, 1998, 37-52 [0003]
• AC McMahon, L. Critharides and HC Lowe: Animal Models of Atherosclerosis Progression: Current Concepts, Curr Drug Targets Cardiovasc Haematol Disord 5, 2005, 433-440 [0003]
• - JC Russell and S. Proctor: Small animal models of cardiovascular disease: tools for the study of the roles of metabolic syndrome, dyslipidemia, and atherosclerosis. Cardiovascular Pathol 15, 2006, 318-330 [0003]
• G. Cinamon and R. Alon: A real-time in vitro assay for studying leukocyte transendothelial migration under physiological flow conditions, J. Immunol. Methods 273 (1-2), 2003, 53-62 [0009]
• - T. Schreiber, V. Shinder et al .: Shear flow-dependent integration of apical and subendothelial chemokines in T-cell transmigration: implications for locomotion and the multistep paradigm, Blood 109 (4), 2007, 1381-1386 [0009]
• - JT Butcher, AM Penrod, AJ Garcia and RM Nerem: Unique morphology and focal adhesion development of valvular endothelial cells in static and fluid flow environments, Arterioscler Thromb Vasc Biol 24, 2004, 1429-1434 [0011]
• - C. Dong, MJ Slattery et al .: In vitro characterization and micromechanics of tumor cell chemotactic protrusion, locomotion, and extravasation, Ann Biomed Eng 30 (3), 2002, 344-55 [0015]
• JJ Chiu, DL Wang et al .: Effects of disturbed flow on endothelial cells, J Biomech Eng 120 (1), 1998, 2-8 [0018]
• JJ Chiu, CN Chen et al .: Analysis of the effect of disturbed flow on monocytic adhesion to endothelial cells, J. Biomech 36 (12), 2003, 1883-95. [0018]
• - Cinamon and Alon (2003) [0018]
• - Schreiber and Shinder et al. (2007) [0018]
• Cinamon and Alon, 2003, and Schreiber and Shinder et al., 2007 [0021]

Claims (52)

Method for detecting the movement and attachment of cells and particles to cell, tissue and implant layers in the simulation of flow conditions, in which the cells and particles in a flow medium through at least one interaction zone in the area of possibly stabilized by support materials cell, tissue and the implantation layer processes are conducted and the movement and attachment processes in laminar, turbulent and pulsating flows are detected visually by monitoring the at least one interaction zone, characterized in that the movement and attachment of the cells and particles, including their migration through the cell, Tissue and implant layer, are optically detected and evaluated in the entire at least one interaction zone, that by the cell, tissue and implant layer transmigrating cells and particles are collected in at least one perfusion chamber preferably below the at least one interaction zone and the flow conditions of the flow medium outside and within the at least one interaction zone, in particular for long-term studies, are controlled by adjusting cell culture parameters such as pH, temperature and nutrient supply to the cells and particles. Process according to claim 1, characterized characterized in that the movement and attachment of the cells and Particles in separate and from several sides on the area of the at least one interaction zone directed observation beam paths is detected. Process according to claim 1, characterized characterized in that the optically detected data of the movement and Attachment of cells and particles digitized and computationally be evaluated. Process according to claim 1, characterized characterized in that the optical detection of movement and attachment the cells and particles are carried out by at least one observation beam path. A method according to claim 4, characterized characterized in that in the at least one observation beam path simultaneously with the observation of the at least one interaction zone a spectroscopic detection of the movement and attachment of the Cells and particles follow. Process according to claim 1, characterized characterized in that the flow conditions as well as the environmental conditions, like pH adjustment, oxygen saturation, the Temperature and nutrient supply for the Cells and particles, adjustable to be regulated. Process according to claim 1, characterized characterized in that the cells and particles for the purpose of their detection by adding one or more fluorescent markers, for example AlexaFluor 488, AlexaFluor 534 or AlexaFluor 647, additional be optically marked. Process according to claim 7, characterized characterized in that the cells and particles by several different Fluorescence marker, additionally optically labeled. Process according to claim 1, characterized characterized in that the cells and particles are for the purpose of their detection, for example, by adding one or more isotopes, in addition be radioactively marked. A method according to claim 1, characterized in that the cell, tissue and implant layers of at least one interaction zone by the addition of one or more fluorescent markers, such as AlexaFluor 488 ^© , AlexaFluor 534 ^© or AlexaFluor 647 ^© or suitable Qdots ^© , optically labeled. Process according to claim 1, characterized characterized in that the cell, tissue and implant layers of the at least one interaction zone, for example by adding a or more isotopes, additionally be radioactively labeled. Process according to claim 1, characterized characterized in that the at least one interaction zone flowing through Flow medium active substances and / or biologically active particles, especially of microbiological origin, such as bacteria, spores, Fungi and parasites, in dead and / or vital form, and / or non-biologically active particles, such as microbeads added become. A method according to claim 12, characterized characterized in that the application of the active substances and / or biologically active particles directly in the at least one Interaction zone takes place. A method according to claim 12, characterized characterized in that the active substances and / or biologically active and / or biologically inert particles with variably dosable Concentration during the detection of movement and attachment of Cells and particles are added. Process according to claim 1, characterized characterized in that the cells and particles are at least one Flow through the interaction zone several times. A method according to claim 1, characterized in that the transmigrating through the cell, tissue and implant layer and in the we at least one of the perfusion chambers collected cells and particles are subjected to a separate additional analysis. Process according to claim 1, characterized characterized in that the pH and the oxygen saturation of the flow medium can be measured by potentiometry. Device for detecting the movement and attachment of cells and particles to cell, tissue and implant layers in the simulation of flow conditions, in which the cells and particles in a flow medium through at least one interaction zone in the possibly stabilized by support materials cell, tissue and implant layer are passed and the movement and attachment processes in laminar, turbulent and pulsating flows are visually detected by supervision of the at least one interaction zone, characterized in that a microscopy device ( 12 ) with at least one observation beam path ( 36 . 37 ) for the optical detection of the cells and particles not only above, but also inside and outside, preferably below, the at least one interaction zone ( 2 . 3 . 4 ), including the detection of the migration of the cells and particles through their cell, tissue and implant layer, it is provided that sensor and control elements (in and / or on the device) ( 28 ) are arranged, through which, in particular for long-term studies, the cell culture parameters of at least one medium cycle ( 26 ) of the device, for example pH, oxygen saturation, temperature and nutrient supply of the cells and particles, can be controlled and adjusted in an adjustable manner, and that preferably below the at least one interaction zone ( 2 . 3 . 4 ) at least one perfusion chamber ( 7 . 8th . 9 . 30 . 31 . 32 ) for introducing test substances into the at least one interaction zone ( 2 . 3 . 4 ) as well as for the enrichment of cells and particles transmigrating through the cell, tissue and implant layer. Device according to claim 18, characterized in that the microscopy device ( 12 ), in addition to the observation beam path which is directed to the at least one interaction zone in a supervisory manner, has an observation beam path directed substantially laterally onto the same interaction zone (US Pat. 36 ) having. Device according to claim 18, characterized in that in addition to the microscopy device ( 12 ) a spectroscopy unit ( 20 . 21 ) for the detection and evaluation of the cells and particles, including the analysis of the at least one interaction zone ( 2 . 3 . 4 ) itself, is provided. Device according to claim 20, characterized in that for detecting the data of the spectroscopy unit ( 20 . 21 ) A laser scanning microscope is provided. Device according to claim 20, characterized in that the spectroscopy unit ( 12 ) for data evaluation with a computer ( 18 ) Device according to claim 19, characterized in that in the observation beam paths ( 36 . 37 ) of the microscopy device ( 12 ) Cameras ( 16 . 17 ) are provided for image acquisition, preferably for image evaluation with a computer ( 18 ) keep in touch. Device according to claim 20, characterized in that the spectroscopy unit ( 20 . 21 ) for the synchronous evaluation of the spectrometric data with the microscopy device ( 12 ) is coupled to the cells and particles in the at least one interaction zone ( 2 . 3 . 4 ) spatially localized. Apparatus according to claim 19, characterized in that the substantially laterally on the at least one interaction zone ( 2 . 3 . 4 ) directed observation beam path ( 36 ) to a separate recording unit ( 17 ) is coupled. Device according to claim 18, characterized in that as a microscopy device ( 12 ) two stereo microscopy systems are provided. Device according to claim 26, characterized in that the two stereomicroscopy systems for synchronous evaluation of the image and / or video data and a as accurate as possible representation of interaction and migration processes the particles and cells in three-dimensional space hard and / or software-coupled with each other. Device according to claim 18, characterized in that means, for example a tripod system, are provided which allow the microscope device or its optical receiving elements to be in position relative to the at least one interaction zone ( 2 . 3 . 4 ) to change. Apparatus according to claim 20, characterized in that means, for example a tripod system, are provided, which allow the spectroscopic unit in position relative to the at least one interaction zone ( 2 . 3 . 4 ) to change. Device according to claim 28 and / or 29, characterized in that said means rail-like guide elements, in particular for the transversal position change along the at least one interaction zone ( 2 . 3 . 4 ) contain. Device according to claim 28 and / or 29, characterized in that these means pivot or rotation elements for changing the angular position to the at least one interaction zone ( 2 . 3 . 4 ) contain. Device according to claim 18, characterized in that, in particular for the purpose of a parallel feasible testing of different active ingredients or different active substance concentrations, preferably below the at least one interaction zone ( 2 . 3 . 4 ) a plurality, for example three, in the flow direction of the cells and particles successively arranged perfusion chambers ( 7 . 8th . 9 . 30 . 31 . 32 ) are arranged. Device according to claim 32, characterized in that the perfusion chambers ( 7 . 8th . 9 . 30 . 31 . 32 ) are constructed and arranged modularly for the purpose of fast and low-cost assembly and / or mechanical conversion. Device according to claim 18, characterized in that in the at least one perfusion chamber ( 7 . 8th . 9 . 30 . 31 . 32 ) Means, for example flow channels, are provided for generating a media swirl, so that transmigrating cells and particles of the flow medium ( 6 ) from the underside of the penetrated cell, tissue and implant layers and in the at least one perfusion chamber ( 7 . 8th . 9 . 30 . 31 . 32 ) are caught. Device according to claim 32, characterized in that in the at least one perfusion chamber ( 7 . 8th . 9 . 30 . 31 . 32 ) Means, for example one or more cell sieves, are provided for collecting the transmigrating cells and particles. Apparatus according to claim 18, characterized in that means are provided, for example a valve system ( 25 ), which allow additional active ingredients in the flow medium ( 6 ) with the cells and particles. Device according to claim 18, characterized in that the cell, tissue and implant layer of the at least one interaction zone ( 2 . 3 . 4 ) is stabilized by one or more optically transparent carrier layers, for example bacterial nanocellulose, for the observation beam paths. Device according to claim 37, characterized in that the cell, tissue and implant layer of the at least one interaction zone ( 2 . 3 . 4 ) consists of several individual cell layers, each sandwiched by one for the observation beam paths ( 36 . 37 ) Optically transparent carrier layer, such as bacterial nanocellulose, are separated. Device according to claim 18, characterized in that means ( 5 . 10 ) for loading, holding and removing the cell, tissue and implant layer into or out of the at least one interaction zone ( 2 . 3 . 4 ) are provided. Device according to claim 39, characterized in that said means move from one to the at least one interaction zone ( 2 . 3 . 4 ) comprise an insertable frame system. Device according to claim 18, characterized in that means are provided, for example a bubble trap, which permits a homogeneous and bubble-free passage of the cells and particles past the cell, tissue and implant layer of the at least one interaction zone ( 2 . 3 . 4 ) guarantee. Device according to claim 18, characterized in that the device is in the form of a circulatory system ( 26 ) in which the cells and particles form the at least one interaction zone ( 2 . 3 . 4 ) flow through several times. Device according to claim 18, characterized in that, in particular for long-term drug tests, reservoirs ( 23 . 24 ) are provided for cells, particles and required nutrient media. Device according to claim 18, characterized in that semipermeable membranes are provided for the exchange of spent and supply of fresh nutrient medium, at which the liquid with the cells, particles and nutrient medium in its circulation ( 26 ) flowed past. Device according to claim 18, characterized in that means ( 11 . 27 . 29 ) are provided for heating the device, for example a heatable reservoir ( 29 ), also from the flow medium with the cells, particles of the flow medium ( 6 ) is flowed through. Apparatus according to claim 18, characterized in that means for CO ₂ admixing of the flow medium ( 6 ) are provided, for example, a nozzle system, the CO ₂ the flow medium ( 6 ) in a reservoir ( 24 ) mixes. Device according to claim 18, characterized in that means ( 28 ) are provided for measuring the pH of the flow medium ( 6 ), for example, ion selective field effect transistors (ISFET) or potentiometric indicator electrodes. Apparatus according to claim 18, characterized in that means are provided for the oxygen admixture of the flow medium, for example an oxygen-gassable reservoir ( 24 ) for the flow medium ( 6 ). Device according to claim 18, characterized in that means ( 28 ) are provided for measuring the pO ₂ value of the flow medium ( 6 ), for example membrane-covered electrodes or electrochemical cells with solid electrolytes. Apparatus according to claim 18, characterized in that means for measuring the flow velocity of the flow medium ( 6 ), for example one in the flow medium ( 6 ) introduced probe with rotary blades. Device according to claim 18, characterized in that means for exciting fluorescent dyes within the chamber body ( 1 ) are provided, for example, a light source, via which excitation light by means of light guide laterally to the chamber body ( 1 ) is introduced. Device according to claim 18, characterized in that means for exciting molecular vibrations in cells or particles above, inside or below the at least one interaction zone ( 2 . 3 . 4 ) are present, for example a laser, the at least one interaction zone ( 2 . 3 . 4 ) and coupled to a confocal microscope.

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