WO2024127090A1

WO2024127090A1 - A device for investigating cell movement

Info

Publication number: WO2024127090A1
Application number: PCT/IB2023/051851
Authority: WO
Inventors: Prafulla Namdeo Shede; Sheetal Popatprasad Pardeshi; Bharat Bajarang Ballal; Ashish Vasant Polkade
Original assignee: Individual
Current assignee: Individual
Priority date: 2022-12-14
Filing date: 2023-02-28
Publication date: 2024-06-20
Anticipated expiration: 2025-06-14

Abstract

A device for investigation of movement of cells under the effect of chemical or other factors comprising peripheral grooves and well connected through a plurality of channels. The elevated plurality of ribs is present in the well forming a central port to channelize the diffusion of chemical or factor. The central port adapted to receive the chemicals under study. The device described in invention can be used for study of chemotaxis, chemoinvasion, phototaxis and other such cell movements. The device will hold the system including the cells and chemicals under investigation so that the cell movements or chemotaxis or chemoinvasion or similar can be analyzed continuously or periodically or as required. The device can be preferably placed in a petri dish or alike or can be operated independently as per requirement.

Description

A DEVICE FOR INVESTIGATING CELL MOVEMENT

TECHNICAL FIELD OF INVENTION

The present invention relates to a device for investigating cell movement. More particularly, the invention relates to a method for investigation of effect of chemical, such as chemo effectors, and other factors, such as light, temperature and others on cell movement.

BACKGROUND OF INVENTION

Various chemotaxis devices have been identified for studying the effect of chemical on the cell movement e.g., Boyden chamber consisting of two chambers, one above the other separated by a membrane wherein the chemo attractant is added to the bottom chamber and cells from the top chamber migrate actively through the membrane in response to the chemical cue. The number of cells migrated are then quantified by traditional cell counting methods. The disadvantages include errors in cell counting; time required for quantification, fragility of membrane used which may tear off during processing and rapid dissipation of the gradient. Further, Zigmond or Dunn chamber are used for the study of directional cell movement which uses videography of cell movement. However, the assay volumes are very small and setting the chamber for gradient is often complex. The agarose assay also has been described where cells are allowed to migrate under the influence of gradient of chemical effector, the gel is then removed and cells fixed using fixating agent like methanol. The fixed cells are then stained and measured optically.

US5302515 discloses a single or multisite chemotaxis chamber with two compartments separated by a thin passage such that cells will have to actively crawl through the passage to reach from first to another chamber in response to chemical gradient. US6238874B1 discloses a single site assay device and multisite high throughput assay device and method is described with two chambers between which the chemical gradient is established and cell motility can be detected using optical system. The system can process multiple samples at a time but is limited to using very small volumes of samples. US6468786B2 has description of cell activity assay apparatus with electromagnetic radiation beam for measurement of cell activity in form of chemotaxis, migration, invasion and others. The apparatus works at low volume and required assay time can be up to 48 hours. US6723523 B2 described a system for study of cell movement under the influence of chemical or other factors by using under-environment system for detecting cell movement by measuring changes in the resistance of the electrode due to cell arrival at the target electrode, that is, by sensing impedance and provides automated multi sample analysis system. US6,921,660 B2 has description of chemotaxis and cell motility device comprising of well regions connected with channel. US6,864,065 B2 discloses an assay for real monitoring of cell motility by preferably using phase contrast microscope or fluorescent image microscope. However, the cost of the device is high and volume used for assay is very low. US7,022,516 B2 discloses amicrowell based apparatus wherein plural number of wells connected by channel with grooves of the dimensions of cells to be studied for detection of chemotaxis and chemotaxis-based separation which requires cell samples in micro quantities and pure or mixed wherein microscopy used for detecting the cell movement.

Disadvantages of prior arts include small volume of sample for analysis, costly devices, complex protocols, inability or difficulty in sterilizing device using autoclaving or other reliable methods, difficulty in maintaining sterility throughout the experiment, cost involved in automated detection. The present application overcomes some of these limitations to provide practically and commercially viable product and process of cell movement investigation.

Thus, there is an unmet need of a cell movement investigation device, method for use of the device which can be applied at wider scale or volume. Also, the ease of maintaining of sterility in the cell movement investigation device is needed. Further, the prior art has failed to provide a device for monitoring chemotaxis and/or chemo invasion, which is not limited to measuring the effects on cell migration of chemo attractants, chemo repellants and chemo stimulants.

Thus, the inventors of the present invention have successfully addressed the existing drawbacks and envisaged a cell movement investigation device which overcomes the above-mentioned limitations and drawbacks.

SUMMARY OF INVENTION

A summary of certain embodiments disclosed herein is set forth below. It should be understood that these aspects are presented merely to provide the reader with a brief summary of these certain embodiments and that these aspects are not intended to limit the scope of this disclosure. Indeed, this disclosure may encompass a variety of aspects that may not be set forth below.

In one aspect, the present disclosure envisages a device for investing the cell movement including a solid base and a well, wherein the well is coaxially located at the center and within the radial axis of the said solid base. The solid base includes a peripheral groove positioned coaxially, adjacent to the distal end of the central axis of the said solid base, wherein said peripheral groove encloses the well; a plurality of channels connecting said well with said peripheral groove; and a plurality of ribs extending orthogonally from the bottom of the well positioned within the radial axis of the said well, the said plurality of ribs is interspaced defining a slot along with a plurality of slit. Furthermore, the rib and the channel are positioned in the solid base, characterized in that the centerline of the slit defined by at least two ribs and centerline of the channel are non-collinear.

In second aspect, the present invention relates to a system for investigating the cell movement comprising a solid base; a well for cell migration and chemical diffusion; a peripheral groove characterized in that the analyte sample is placed at least one region within the peripheral groove, wherein the peripheral groove links at least two channel; a plurality of channels connecting said well with said peripheral groove characterized in that cell migration and chemical diffusion occurs at channels; a plurality of ribs defining a slot and plurality of slit characterized in that the ribs acts as barrier and channelize at least one chemical and cell effector through the slit and avoid free flow of fluid medium; wherein the slit acts as a passage for movement of cell, diffusion of chemical and cell effectors, wherein slot holds at least one selected from chemical and cell effectors; and a wall characterized in that the wall acts as a barrier for holding at least one solid, semisolid, liquid and combinations thereof.

In third aspect, the present invention relates to a method for investigating the cell movement comprising a) adding a buffer solution of predetermined quantity on at least a portion on the peripheral groove; b) adding an analyte on at least one portion on the peripheral groove, bisecting at least the two channels; c) adding at least a cell effector at portion of the slot in the well region; and d) collecting the sample of predetermined quantity from at least a portion defined channel connecting the opening of the well.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention will now be explained in more detail with reference to the following exemplary non-limiting embodiments and the accompanying drawings, in which:

Figure 1 A shows a top view of a portion of an embodiment of the test device according to the present invention.

Figure IB shows a side elevational view of a longitudinal cross section, of an embodiment of the test device according to the present invention.

Figure 2 shows a top perspective view of an embodiment of test device according to the present invention. Figure 3 shows a process flow diagram of a method according to an embodiment of the present invention.

DETAILED DESCRIPTION OF THE EMBODIMENTS

The present invention relates to a device for investigating effect of chemical or other factors on cell movement and to the construction or making of such a device. Under the effect of chemicals here means the positive, negative, neutral or any effect on cell movement in presence of any chemical at a given concentration or gradient of the chemical or absence of the chemical. Other factor here may include a temperature, pH, light, smell or any other physical factor that may influence the cell movement in positive, negative, neutral or any other way. Further, the present invention also relates to the process of using said device for the inference of cell movement under the effect of abovementioned chemicals or factors.

In one embodiment of the present invention the device wherein different samples containing cells of pure or plural types can be screened for movement in response to such chemicals or factors of which a gradient is created in the device. The device in the present invention further aims at providing a system to enrich/ enhance/ augment such cells that respond in the form of cell movement to chemicals or factors under study. The invention further aims to provide a system where samples of different origins can be used for foregoing and ensuing purposes mentioned. Furthermore, the present invention aims at providing a device by which investigation of cells can be done by “movement based on their own action”. “Movement based on their own action” here means cell movement can be investigated without, for example, cells have to deform or experience, for example, pressure.

In one embodiment of the present invention, the device for investing phenomenon like chemotaxis where a laminar gradient of diffusible chemical can be created in the device and the device can be closed to avoid interference of external environment. The movement of cells can be then investigated under the influence of this gradient in terms of positive chemotaxis, that is, movement of cells towards the increasing gradient of chemical or negative chemotaxis, that is, movement of cells away from the increasing gradient of chemical or neutral or other effect.

In one embodiment of the present invention the device can be used at higher volumes, of samples contOaining single type or plural types of cells which can appropriately represent the sample. The device described in present invention can further be used in different sizes of the devices as per the requirement of volume of sample is to be processed that can be sufficient to achieve the objectives of investigation, such as, dose response, cell movement rate, enrichment and others. The device in the present invention can further be used for the foregoing and ensuing purposes for continuous or periodic observations of the object under investigation.

As shown in Figure 1A, according to one embodiment of the present invention, a test device 100, such as, for example, chemo taxis, photo taxis, thermo taxis, chemo invasion device, includes a solid base 10 and a well 20, wherein the well 20 is coaxially located at the center and within the radial axis of the said solid base 10. In one embodiment, at least the diameter of the well is less than diameter of the solid base. In preferred embodiment, the diameter of the well 20 is less than radius of the solid base. The solid base 10 includes a peripheral groove 30 positioned coaxially, adjacent to the distal end of the central axis of the said solid base, wherein said peripheral groove 30 encloses the well 20. Further, the said well 20 and the peripheral groove 30 is connected by at least a plurality of channels 40. The well 20 and the peripheral groove 30 are configured such that they together define a plurality of channels 40 as shown. Preferably test device 10 comprises a plurality of channels, as shown by way of example in the embodiment of Figure 1A. In one embodiment, at least the plurality of channels connecting the well and peripheral grooves 30 are placed equiangular. In preferred embodiment, the device includes four channels connecting the well 20 and peripheral grove 30. In one embodiment, the channel 30 originates from the interior wall of the peripheral groove 32 to at least one opening at the wall of the well 20. In a preferred embodiment, the channel 40 is linear connecting well 20 and peripheral groove 30. The plurality of channels 40 allow the flow of analyte and fluid medium from well to the peripheral grove 30 and vice versa in a test orientation of the device. The "test orientation" of the device is meant to refer to a spatial orientation of the device during testing. As seen in Figure IB, the well is defined by a through-hole in solid base, corresponding to well 20, and by an upper surface U of solid base 10. In particular, the sides of the well 20 is defined by the walls of the through holes in the solid base 10, and the bottoms of well is defined by the upper surface U of solid base 10. It is noted that in the context of the present invention, "top," "bottom," "upper" and "side" are defined relative to the test orientation of the device. As seen collectively in Figures 1A, IB and 2, a length L of channel region 40 is defined in a direction of the longitudinal axis of channel region 40; a depth D of channel region 40 and peripheral groove 30are defined in a direction normal to upper surface U of solid base 10; a width W is defined in a direction normal to the length L and depth D of channel region 40. The width of the peripheral groove region 30 is defined in a direction normal to the depth D of peripheral groove region 30. In one embodiment, the bottom of well 20, channel 40 and peripheral groove 30 are coplanar.

In one embodiment, wherein the exterior portion of the peripheral groove 31 defines a wall 70 surrounding the peripheral groove 30. The wall forms an outer barrier wall of the device 100.

Regarding the shape of device, the device is not limited to hexagonal, cylindrical, cuboidal, circular, ellipsoidal, rectangular, square, or any other polygonal or curved shape.

Regarding the dimensions of a channel 40, the length L of a given channel 40 can vary based on various testing parameters. For instance, the length L of a given channel 40 may vary in relation to the distance over which cell movement e.g., chemotaxis is required to occur. For example, the length L of a given channel 40 can range from about 10mm to about 20mm in order to allow cells a sufficient distance to travel and therefore sufficient opportunity to observe cell chemotaxis and chemoinvasion. The width W and depth D of a given channel 40 may also vary as a function of various test parameters. For examples, the width W and depth D of a given channel 40 may vary, in a chemotaxis, and/or chemo invasion device, depending on the size of the cell being studied. Generally, where the test device is a chemotaxis, haptotaxis and/or chemoinvasion device, a given channel 40’ s width W and depth D may be approximately in the range of the diameter of the cell being assayed.

In another embodiment of the present invention the device further comprises a plurality of ribs 50 extending orthogonally from the bottom of the well positioned within the radial axis of the well. The plurality of ribs is mounted to the bottom of the well 20 by being placed in substantially fluid-tight, conformal contact with the bottom of the well 20. In the context of the present invention, "conformal contact" means substantially form-fitting, substantially fluid-tight contact. The plurality of ribs 50 is adapted to hold the chemical or factor under investigation and is preferably located in front of the connector channel 40 to guide the flow, for example, diffusion of chemical. In one embodiment, the plurality of ribs takes the shape of an arc which is interspaced defining a slot at the center of the well 20 and a plurality of slits 60. In one embodiment, the height of the plurality of ribs is at least equal to or less than the depth of the well, preferably less than the depth of the well 20. In another preferred embodiment, the numbers of ribs are equal to the number of channels 40.

Furthermore, the rib 50 and the channel 40 are positioned in the solid base, in such a position that the centerline of the slit 60 defined by at least two ribs 50 and centerline along the length of the channel are non-collinear. In one embodiment, the ribs are positioned in any spatial configuration within the well and concentric within the well. In preferred embodiment, the ribs is positioned within the well such that line bisecting the width of the channel 40 is equal to the center of the rib arc. The configuration of the ribs 50 forms a central port to be adapted to hold the chemical or factor under investigation and is preferably located in front of the connector channel 40 to guide the flow, for example, diffusion of chemical. In an alternative embodiment, the device doesn’t include a plurality of ribs.

In a preferred embodiment, the device has circular shape having an outer diameter in the range of 21 to 210 mm. The peripheral groove is circular in shape having an outer diameter in a range 20-200 mm. The height of the plurality of ribs in a range of 0.1-30 mm.

In one embodiment, the solid base is prepared from material selected from at least one but not limited to acrylic, teflon, stainless steel, polypropylene, glass, metals and combinations thereof.

The size of these embodiments may differ depending on the size of the planar base on which these embodiments are supported. The optimum dimensions of channels and diameters of device depend upon the type of cells being studied. The device my take any shape and the shape of embodiments may also differ with respect to each other. The device may preferably be assembled in petri dish or alike with a lid or can be used separately.

According to one aspect, the invention describes a system for investigating of the cell movement comprising a solid base 10; a well 20 for cell migration and chemical diffusion; a peripheral groove 30 characterized in that the analyte sample is placed at least one region within the peripheral groove 30, wherein the peripheral groove 30 links at least two channel 40; a plurality of channels 40 connecting said well 20 with said peripheral groove 30 characterized in that cell migration and chemical diffusion occurs at channels 40;a plurality of ribs 50 defining a slot 80 and plurality of slit 60 characterized in that the ribs acts as barrier and channelize at least one chemical and cell effector through the slit 60 and avoid free flow of fluid medium; wherein the slit 60 acts as a passage for movement of cell, diffusion of chemical and cell effectors, wherein slot 80 holds at least one selected from chemical and cell effectors; and a wall 70 characterized in that the wall acts as a barrier for holding at least one solid, semisolid, liquid and combinations thereof.

According to one aspect, the invention describes a method for investigating of the cell movement including as disclosed in Fig no. 3, a buffer solution of predetermined quantity is added on at least a portion 310 on the peripheral groove 30. Preferably, the buffer solution is added till the level of at least depth of the channel 40.

In one embodiment, an analyteis added on at least one portion 320 in the peripheral groove 30, wherein the portion 320 is located on a line bisecting at least the two channels 40; the analyte sample can be added at suitable quantity to study the effect on the biological samples.

In one embodiment, a cell effector is added at the lot 80 of the well region 330. The cell effector is at least one selected from chemical, light, temperature, pH and smell; and collecting the sample of predetermined quantity from at least a portion 340 defined channel connecting the opening of the well 20.

In another embodiment, the predetermined quantity of analyte sample is selected from range 0.01 ml to 0.5 ml.

In one embodiment of the present invention, the peripheral groove 30 is adapted to receive a buffer solution and an analyte. Comprising at least one cell selected from prokaryotes and eukaryotes.

In one embodiment of the present invention the device that can be used separately, preferably covered with a suitable detachable lid or can be used in a system like Petri dish available in different dimensions wherein the shape of device can be circular and size of the device can be different depending on the size of Petri dish. Moreover, the shape of the device described in present invention can also be substantially semicircular, rectangular, square, polygonal or other. The present invention further aims at providing a device that can be sterilized by different means, such as autoclaving, and can be used in aseptic operations as required. The device can be used without sterilization if the requirement be.

From the foregoing, it will be observed that numerous modifications and variations can be effected without departing from the true spirit and scope of the novel concept of the present invention. For example, different embodiments of a device of the present invention may be combined. Embodiments of the present invention further contemplate different types of assays, for example, an assay wherein the test agent comprises a buffer solution instead of a chemotactic agent. In such an assay, cell migration through channel region 30 is observed in the absence of a chemotactic gradient.

It will be appreciated that the present disclosure is intended to set forth the exemplifications of the invention, and the exemplifications set forth are not intended to limit the invention to the specific embodiments illustrated. The disclosure is intended to cover by the appended claims all such modifications as fall within the spirit and scope of the claims.

Examples hereinafter, although an Example demonstrates this invention further in detail, this invention is not limited to these.

Experiment 1

The process to use the device e.g., chemotaxis assay plate includes placing a cellulose thimble having outer diameter same as that of the internal diameter of the central port. The thimble of outer diameter 11 mm, made of what man paper grade 1 was used. This assembly was then placed in petri dish of optimum size, if required. The chemotaxis assay plate of outer diameter of 88 mm and placed the assay plate in petri dish of outer diameter of 90 mm and height 12 mm. The autoclaved the assembly at 121° C, 15 psi pressure, for 15 minutes. The assembly was placed on coplanar platform without any slope to create perfect gradient of chemical under study. 13.05 ml of chemotaxis buffer of 100 mM and pH 7 was added to device followed by 0.45 ml of 100 mM L-aspartate as chemoeffector. The cells used for optimization of the process were purchased from National Centre for Microbial Resources (NCMR), Pune, previously known as Microbial Culture Collection (MCC), Pune. The Culture Collection ID of the cells used in the described process is MCC 2049, Bacillus subtilis and MCC 2989, Pseudomonas putida. The initial concentration of cells used for assay were 10⁷- 10⁸ cells which were grown overnight in nutrient broth and harvested by centrifugation at 6000 rpm at 25° C for 10 minutes. The cells were washed with abovementioned buffer to remove media components. 375 pL of cell suspension of optical density 1 at 620 nm was added at all sample addition points as illustrated in accompanying workflow diagram in Fig. 3 giving total volume of cell sample 1.5 ml. In the control experiment, chemical or factor was replaced with buffer only or other suitable neutral factors. 0.45 ml of chemotaxis buffer instead of L-aspartate in control experiment in the process. After addition at the central port, samples was withdrawn at regular time intervals as per requirement from any or all of the sample collection areas 340. The 0.1 ml of sample from one of the sample collection area at the intervals of 5 to 10 minutes was used for analysis. The samples withdrawn from the chemotaxis assay plate was then analyzed using suitable methods of analysis, such as, total viable count for microbial cells. The count of test experiment was compared with that of the control experiment to interpret the results.

Experiment 2

The process to use the device e.g., chemotaxis assay plate includes placing a cellulose thimble having outer diameter same as that of the internal diameter of the central port. The thimble of outer diameter 11 mm, made of Whatman paper grade 1 was used. This assembly was then placed in petri dish of optimum size, if required. The chemotaxis assay plate of outer diameter of 88 mm and placed the assay plate in petri dish of outer diameter of 90 mm and height 12 mm. The autoclaved the assembly at 121° C, 15 psi pressure, for 15 minutes. The different combination of components in experiment as follows. 13.2 ml of Phosphate buffer of 100 mM and pH 7 was added to device followed by 375 pL of cell suspension as described and processed in example 1 at all sample addition points. The chemoeffector, 0.3 ml of 5 mM L-aspartate was added in the thimble. In the control experiment, chemical or factor was replaced with buffer only or other suitable neutral factors. 0.3 ml of chemotaxis buffer in thimble instead of L- aspartate in control experiment in the process. The withdrawal of samples from device and its processing was done as described in example 1.

Experiment 3

The workflow of the device for best method is as illustrated Fig. no. 3 of workflow diagram and described in following section. The process to use the chemotaxis assay plate includes placing a cellulose thimble having outer diameter same as that of the internal diameter of the central port. The thimble of outer diameter 11 mm, made of Whatman paper grade 1. This assembly was placed in petri dish of optimum size, if required. The chemotaxis assay plate (made of Teflon material) of outer diameter of 88 mm and placed the assay plate in petri dish of outer diameter of 90 mm and height 15 mm. This assembly was autoclaved at 121 °C, 15 psi pressure, for 15 minutes. As illustrated in the accompanying workflow Fig. no 3, the step 1, is addition of buffer, preferably chemotaxis buffer having molarity of 10 to 100 mM and pH 7. The quantity used by inventors in the optimized process was 14.75 ml of chemotaxis buffer of the composition. Step 2 of the process was addition of sample containing cells to be investigated for cell movement. The sample containing desired density of single or plural type of cells was added in required concentration to give desired effective dilution. 37.5 microliters of sample were added at all sample addition points as illustrated in accompanying workflow Fig. no 3. The total volume of sample thus achieved was 0.15 ml. The cells used for optimization of the process were purchased from National Centre for Microbial Resources (NCMR), Pune, previously known as Microbial Culture Collection (MCC), Pune. The Culture Collection ID of the cells used in the described process are MCC 2049, Bacillus subtilis and MCC 2989, Pseudomonas putida. The initial concentration of cells used for assay were 10⁷- 10⁸ cells. 0.15 ml of said density culture was added to 14.75 ml of buffer in chemotaxis assay plate as shown in workflow diagram in Fig. no 3, to give 100-fold dilution of the cell density. The cultures were added slowly using micropipette, such that minimum ripples were introduced in the buffer in chemotaxis assay plate. In the test experiment, step 3 of the process is to add chemical or factor under study at required concentration at the central port. L-aspartate, a known chemoeffector for cells described previously was used as chemoeffector. 0.1 ml of 3 mM L-aspartate solution at pH 7 was added in the cellulose thimble placed at the central port as described in the beginning of this section and illustrated in the workflow diagram. Whereas, in the control experiment, chemical or factor should be replaced with buffer only or other suitable neutral factors. 0.1 ml of chemotaxis buffer instead of L-aspartate in control experiment in the process described in this document. After addition at the central port, samples can be withdrawn at regular time intervals as per requirement from any or all of the sample collection areas as shown in workflow Fig. no 3. 0.1 ml of sample from one of the sample collection area at the intervals of 5 to 10 minutes and used for analysis. The samples withdrawn from the chemotaxis assay plate was analyzed using suitable methods of analysis, such as, total viable count for microbial cells. The count of test experiment was compared with that of the control experiment to interpret the results. The process was adopted to study any chemical attractant at optimum concentration for chemotaxis by any prokaryotic cells, pure, mixed or environmental sample, as described above and for eukaryotic cells, for example, lymphocytes, by using optimum chemical concentration and initial cell density.

The principle of above method discloses in Examples 1-3 is performed by creating a gradient of the chemical or factor under study, to which the cells respond in terms of movement towards or away from the increasing gradient of the chemical or factor and the movement can be inferred by comparing with control experiment.10³ to 10⁴ fold more cells of MCC 2049, Bacillus subtilis and MCC 2989, Pseudomonas putida were recovered in comparison to control as depicted in Table 1 and Table 2 mentioned below. Table 1 : Chemotaxis study using 3mM Aspartate as chemoattractant and Pseudomonas putida MCC 2989 as a test organism

TNTC: Too numerous to count Table 2: Chemotaxis study using 3mM Aspartate as chemoattractant and Bacillus subtilis MCC 2049 as a test organism

TNTC: Too numerous to count Further, the device described in the invention can also be used for study of phototaxis by providing a photon source through the lid. Further, the abovementioned purpose, the device should be placed in dark, that is, a box or dark room or black colored case or alike. The photon source to be studied is placed such that the position of photon source is exactly above the central port. Rest of the protocol can be followed in a similar manner as described for study of chemotaxis in the preceding section.

The device described in the invention can also be used to study thermotaxis, wherein a heating probe can be inserted at the central port and a gradient of temperature can created in the system. Rest of the protocol can be followed in a similar manner as described for study of chemotaxis in the preceding section. The device can also be adopted to study chemoinvasion by tumor cells by using a ring of filter paper of appropriate pore size (as per cells under study) coated with optimum thickness of commercially available matrix gel of similar gels as per requirement. The coated ring can be placed at the opening of connecting channel into to inner channel. The cells to be studied for chemoinvasion can be added at the connecting channels near the filter paper ring. The chemical attractant to be used as per requirement is added at the central port and the cells invading through the filter paper can be collected at the inner channel. The analysis of collected cells can be done using previously described methods such as microscopic, spectrophotometric, colorimetric or other suitable methods.

Benefits over prior art

The present invention can provide following benefits or advantages over prior art-

1. Reduced complexity of process to study response of cells in terms of motility on them on to the gradients or factors under study.

2. More volume of sample can be used for analysis of the above stated response.

3. Single sampling or intermittent sampling at various time intervals can be done from the same experiments to provide time course study as per requirement. 4. The optimization of time of experiment for best result can be done and the time required can be less as compared to previous processes described.

5. The device described in invention can be used to study the said response using liquid or semi-solid or gel support medium. 6. The design of the device described in present invention is simple and can be taken for commercial production easily.

7. a cost effective and reusable tool for investigation of cell migration or movement under the effect of chemicals or other factors.

8. Mixed cell populations can be studied.

Claims

CLAIM

1. A device 100 for investigating the cell movement comprising: a solid base 10; a well 20 located at the center and within the radial axis of the said solid base 10, wherein said well 20 is coaxial with the solid base 10; a peripheral groove 30 positioned coaxially, adjacent to the distal end of the central axis of the said solid base 10, wherein said peripheral groove 30 enclosing the well 20; and a plurality of channels 40 connecting said well 20 with said peripheral groove 30.

2. The device as claimed in claim 1 , wherein the device further comprises a plurality of ribs 50 extending orthogonally from the bottom of the well 20 positioned within the radial axis of the said well 20, the said plurality of ribs is interspaced defining a slot 80 along with a plurality of slit 60.

3. The device as claimed in claim 1, wherein the rib 50 and the channel 40 are positioned in the solid base 10, characterized in that the centerline of the slit 60 defined by at least two ribs 50 and centerline of the channel 40 are non-collinear.

4. The device as claimed in claim 1, wherein the shape of said solid base is at least one selected from to hexagonal, cylindrical, cuboidal, circular, ellipsoidal, rectangular and square.

5. The device as claimed in claim 1, wherein the bottom of well 20, channel 40 and peripheral groove 30 are coplanar.

6. The device as claimed in claim 1, wherein the peripheral groove 30 links at least two channels 40. The device as claimed in claim 1, wherein the exterior portion of the peripheral groove 31 defines a wall 70 surrounding the peripheral groove 30. The device as claimed in claim 1, wherein the height of the peripheral groove 30 is less than height of solid base 10. The device as claimed in claim 1, wherein the channel 30 originates from the interior wall of the peripheral groove 32 to at least one opening at the wall of the well 20. The device as claimed in claim 1, wherein the height of the ribs 50 is less than height of solid base 10. The device as claimed in claim 1, wherein the solid base material at least one selected from acrylic, teflon, stainless steel, polypropylene, glass, metals and combinations thereof. A system for investigating the cell movement comprising: a solid base 10; a well 20 for cell migration and chemical diffusion; a peripheral groove 30 characterized in that the analyte sample is placed at least one region within the peripheral groove 30, wherein the peripheral groove 30 links at least two channel 40 a plurality of channels 40 connecting said well 20 with said peripheral groove 30 characterized in that cell migration and chemical diffusion occurs at channels 40; a plurality of ribs 50 defining a slot 80 and plurality of slit 60 characterized in that the ribs acts as barrier and channelize at least one chemical and cell effector through the slit 60 and avoid free flow of fluid medium; wherein the slit 60 acts as a passage for movement of cell, diffusion of chemical and cell effectors, wherein slot 80 holds at least one selected from chemical and cell effectors; a wall 70 characterized in that the wall acts as a barrier for holding at least one solid, semisolid, liquid and combinations thereof. A method for investigating the cell movement using a comprising a) adding a buffer solution of predetermined quantity on at least a portion 310 on the peripheral groove 30; b) adding an analyte on at least one portion 320 in the peripheral groove 30, wherein the portion 320 is located on a line bisecting at least the two channels 40; c) adding at least a cell effector at portion 330 of the slot 80; and d) collecting the sample of predetermined quantity from at least a portion 340 defined channel connecting the opening of the well 20. The method as claimed in claim 13, wherein the analyte is a cell at least one selected from prokaryotes and eukaryotes. The method as claimed in claim 13, wherein the cell effector is at least one selected from chemical, light, temperature, pH and smell.