Surface-enhanced raman scattering-based drug reactivity test method

Abstract

In a surface-enhanced Raman scattering-based method for testing drug reactivity of cerebrovascular cells, a SERS substrate is manufactured. A SERS substrate array including a plurality of the SERS substrates is manufactured. Cells are cultured on the SERS substrate array. A drug is administered to the cultured cells. A Raman signal is measured from the cells to which the drug is administered. An optimal drug is selected from among the administered drugs on the basis of the measured Raman signal.

Description

Surface-enhanced Raman scattering-based drug responsiveness testing method

[0001] The present invention relates to a method for testing drug responsiveness, and more specifically, to a substrate surface-enhanced Raman scattering-based drug responsiveness testing method in which a subject for responsiveness testing is cultured on a SERS (surface-enhanced Raman scattering) substrate array and responsiveness to various drug administrations is tested using Raman signals.

[0002] Laban spectroscopy is applied as the most sensitive chemical analysis method capable of determining the structure and characteristics of molecules by exhibiting distinct signals depending on the molecular structural characteristics of the sample; this method is widely utilized not only in the field of chemistry but also in the analysis of cells and infectious agents.

[0003] For example, Korean Registered Patent No. 10-2225543 discloses a technology using the above-mentioned Raman spectroscopy in exhalation-based lung cancer diagnosis, and Japanese Registered Patent No. 7291112 discloses a technology for performing analysis of myocardial cells using the above-mentioned Raman spectroscopy.

[0004] In particular, to perform analysis using such Raman spectroscopy, it is necessary to fabricate a surface-enhanced Raman scattering (SERS) substrate that significantly amplifies the Raman signal; generally, such SERS substrates must be fabricated to include nanostructures made of precious metal materials.

[0005] However, conventionally, such SERS substrates were merely fabricated as single substrates, which presented limitations when culturing cells or performing repeated experiments, and thus the development of related technologies is required.

[0006] Accordingly, the technical problem of the present invention is conceived from this point, and the objective of the present invention is to provide a substrate surface-enhanced Raman scattering-based drug responsiveness testing method that tests the responsiveness to various drug administrations using Raman signals by culturing a subject for responsiveness testing on a SERS (surface-enhanced Raman scattering) substrate array.

[0007] In a method for testing drug responsiveness according to an embodiment for realizing the objective of the present invention described above, a SERS substrate is fabricated. A SERS substrate array comprising a plurality of said SERS substrates is fabricated. Cells are cultured on said SERS substrate array. A drug is administered to said cultured cells. Raman signals are measured from said cells to which the drug has been administered. Based on said measured Raman signals, an optimal drug among said administered drugs is selected.

[0008] In one embodiment, the cell is a rat brain endothelial cell, and the optimal drug may be a drug with the best reactivity to the cell.

[0009] In one embodiment, in the step of selecting the optimal drug, if a new peak is identified in the measured Raman signal or the size of an existing peak changes, it can be determined that the responsiveness to the cell is excellent.

[0010] In one embodiment, the reactivity to the cell may be a characteristic that allows the cell to pass through a barrier.

[0011] In one embodiment, in the step of fabricating the SERS substrate, nanoparticles can be sputtered onto a base substrate to deposit the nanoparticles on the base substrate.

[0012] In one embodiment, the nanoparticles can pass through the pores of the porous substrate and be sputtered onto the base substrate.

[0013] In one embodiment, the size of the porous portion is larger than the nanoparticle, and the arrangement of the porous portion may be a random arrangement.

[0014] In one embodiment, in the step of fabricating the SERS substrate array, the SERS substrates can be aligned in a predetermined number of columns and rows to fabricate the SERS substrate array on a single plate.

[0015] In one embodiment, the step of culturing the cells may include thawing frozen cells, centrifuging the thawed cells, adding a medium to the centrifuged cells to prepare a cell solution, providing the cell solution to the SERS substrate array, and culturing the cells while replacing the medium.

[0016] In one embodiment, in the step of centrifuging the thawed cells, a medium may be added to the thawed cells and centrifuged to remove the supernatant.

[0017] In one embodiment, in the step of providing the cell solution, the cell solution can be provided to precipitate the SERS substrates.

[0018] In one embodiment, the step of measuring the Raman signal may include moving the SERS substrate array containing the cultured cells to which the drug has been administered to a Petri dish, removing the medium, moving the SERS substrate array to a measurement plate, optimizing the Raman signal measurement conditions of the Raman spectrometer, and measuring the Raman signal under the optimized measurement conditions.

[0019] In one embodiment, the step of measuring the Raman signal can be performed repeatedly at preset intervals while the drug is administered.

[0020] In one embodiment, in the step of optimizing the Raman signal measurement conditions, the distance between the Raman spectrometer and the SERS substrate array can be varied.

[0021] In one embodiment, the optimized Raman signal measurement condition may be the distance between the Raman spectrometer and the SERS substrate array when the R6G (Rhodamine 6G) signal is most clearly and significantly produced.

[0022] According to embodiments of the present invention, a SERS substrate array is fabricated to include a plurality of SERS substrates, allowing for simultaneous cell culture and the simultaneous determination of the drug effects of each drug by administering different drugs to each cultured cell, thereby improving the speed and relative comparative performance of drug responsiveness testing.

[0023] In particular, regarding the examination of the above-mentioned drug responsiveness, for example, it has been verified that the examination can be performed based on Raman signals targeting mouse brain vascular cells, and thus the functionality of the blood-brain barrier of mouse brain vascular cells can also be confirmed through Raman signals.

[0024] In addition, when fabricating the above SERS substrate, instead of growing nanoparticles directly on the substrate, nanoparticles are deposited on the substrate through sputtering to pass through the porous structure, thereby reducing the time required for nanoparticle deposition and enabling the fabrication of a more rapid and effective SERS substrate.

[0025] In addition, by optimizing the measurement conditions of the Raman signal before confirming the responsiveness of the cell by measuring the Raman signal, the accuracy of the signal can be further improved when confirming the responsiveness of the cell solely through the Raman signal.

[0026] FIG. 1 is a flowchart illustrating a drug reactivity test method according to one embodiment of the present invention.

[0027] FIG. 2a is a schematic diagram illustrating the steps for fabricating the SERS substrate of FIG. 1, FIG. 2b is a schematic diagram illustrating an example of the SERS substrate fabricated through FIG. 2a, and FIG. 2c is a schematic diagram illustrating an example of a SERS substrate array.

[0028] Figure 3 is a flowchart illustrating the step of culturing cells to be tested on the SERS substrate array of Figure 1.

[0029] Figure 4 is a schematic diagram illustrating an example of the culture of Figure 3.

[0030] Figure 5 is a flowchart illustrating the steps for measuring the Raman signal of Figure 1.

[0031] Figure 6 is a graph illustrating the steps for optimizing the Raman signal measurement conditions of Figure 5.

[0032] Figures 7a to 7f are graphs showing the results of measuring Raman signals during the culture process of the cells under examination.

[0033] <Explanation of Symbols>

[0034] 100: Base substrate 200: Particle supply unit

[0035] 210 : Porous substrate 211 : Porous area

[0036] 300 : Nanoparticles 310 : Layered particles

[0037] 400: SERS substrate 401: SERS substrate array

[0038] 500 : Cultured cells

[0039] The present invention is susceptible to various modifications and may take various forms, and embodiments are to be described in detail in the text. However, this is not intended to limit the invention to the specific disclosed forms, and it should be understood that the invention includes all modifications, equivalents, and substitutions that fall within the spirit and scope of the invention. Similar reference numerals have been used for similar components in the description of each figure. Terms such as "first," "second," etc., may be used to describe various components, but said components should not be limited by said terms.

[0040] The above terms are used solely for the purpose of distinguishing one component from another. The terms used in this application are used merely to describe specific embodiments and are not intended to limit the invention. The singular expression includes the plural expression unless the context clearly indicates otherwise. In this application, terms such as "comprising" or "consisting of" are intended to specify the existence of the features, numbers, steps, actions, components, parts, or combinations thereof described in the specification, and should be understood as not precluding the existence or addition of one or more other features, numbers, steps, actions, components, parts, or combinations thereof.

[0041] Hereinafter, preferred embodiments of the present invention will be described in more detail with reference to the attached drawings.

[0042] FIG. 1 is a flowchart illustrating a drug reactivity test method according to one embodiment of the present invention.

[0043] Referring to FIG. 1, the drug responsiveness test method according to the present embodiment (hereinafter, test method) is a method of culturing cells to be subject to a responsiveness test, administering various types of drugs to the cultured cells, and testing how each administered drug performs a reaction with respect to the cells.

[0044] At this time, the cells being cultured, that is, the cells for determining reactivity, are not limited to specific cells, but may, for example, be rat brain endothelial cells. Furthermore, the drug is a drug that passes through the cell barrier of the rat brain endothelial cells, and the drug with the best reactivity corresponds to the drug that passes through the rat brain endothelial cells most effectively. In particular, a drug that passes through the rat brain endothelial cells effectively as described above corresponds to a candidate group of drugs capable of treating diseases such as dementia; therefore, the testing method according to this embodiment is a testing method that performs performance evaluation or testing of a drug using the so-called Raman enhancement method, which can ultimately serve as the basis for a method to treat diseases such as human dementia. Of course, if the cells subject to testing are other cells other than rat brain endothelial cells, it is self-evident that the drug corresponds to a drug that passes through the cell barrier of such cells.

[0045] More specifically, the above inspection method is explained as follows.

[0046] FIG. 2a is a schematic diagram illustrating the steps for fabricating the SERS substrate of FIG. 1, FIG. 2b is a schematic diagram illustrating an example of the SERS substrate fabricated through FIG. 2a, and FIG. 2c is a schematic diagram illustrating an example of a SERS substrate array.

[0047] First, referring to FIGS. 1 and FIG. 2a, in the inspection method above, a surface-enhanced Raman scattering (SERS) substrate is fabricated (step S10).

[0048] More specifically, in the step of manufacturing the SERS substrate (step S10), nanoparticles (300) are sputtered onto a base substrate (100) to deposit nanoparticles (310) on the base substrate (100) to manufacture the SERS substrate (400).

[0049] At this time, the nanoparticles (300) are provided through a particle providing unit (200) provided on the upper part of the base substrate (100), and the particle providing unit (200) has a separate porous substrate (210) on the lower part. As illustrated, the porous substrate (210) is a substrate having a porous portion (211) formed therein, and the nanoparticles (300) provided by the particle providing unit (200) pass through the porous portion (211) of the porous substrate (210) and are sputtered onto the base substrate (100) located on the lower part.

[0050] At this time, although the drawings show that the porous portions (211) are arranged at regular intervals, the arrangement of the porous portions (211) can be varied in many ways and may have a random arrangement. In addition, it is obvious that the processing portion (211) must be formed larger than the size of the nanoparticle (300) provided by the particle providing portion (200).

[0051] Thus, as the nanoparticles (300) are sputtered onto the base substrate (100), a stacked particle (310) in which nanoparticles are stacked is formed on the upper surface of the base substrate (100), thereby producing the SERS substrate (400) in the form of a stacked particle (310). In addition, the form in which the nanoparticles (300) are stacked can be any form as illustrated, and as the nanoparticles (300) are stacked on the base substrate (100), they can be formed to a predetermined height.

[0052] At this time, the nanoparticles, i.e., the stacked particles (310), may include a precious metal material such as gold or silver.

[0053] Meanwhile, as shown in FIG. 2b, a plurality of the above SERS substrates are manufactured.

[0054] Afterwards, referring to FIGS. 1 and FIG. 2c, a SERS substrate array (401) comprising a plurality of fabricated SERS substrates (400) is fabricated (step S20).

[0055] That is, as shown in FIG. 2a, individual SER substrates (400) produced by sputtering nanoparticles are arranged in a predetermined number of columns and rows to produce the SERS substrate array (401). At this time, the number of SERS substrates included in the SERS substrate array (401), the number of columns and rows of the SERS substrates, or the spacing can be varied.

[0056] Afterward, referring to FIG. 1, the cells are cultured on the fabricated SERS substrate array (401) (step S30). At this time, the cells being cultured are described as rat brain endothelial cells as an example, as previously described, but are not limited thereto.

[0057] The step of culturing the above-mentioned cells (step S30) is described in more detail as follows.

[0058] FIG. 3 is a flowchart illustrating the step of culturing cells to be examined on the SERS substrate array of FIG. 1. FIG. 4 is an image illustrating an example of the cultivation of FIG. 3.

[0059] That is, referring to FIG. 3, in the step of culturing the cells (step S30), the cells stored in a frozen state are first thawed (step S31). In this cell thawing step, the surface of the storage container in which the cells are stored is first washed with, for example, 70% concentration ethanol (EtOH), and the cells are thawed by placing a medium heated to about 37°C, which is similar to human body temperature.

[0060] In this case, the above-mentioned medium may be, for example, a commercially available product such as IMDM (Iscove's Modified Dulbecco's Medium).

[0061] Afterward, referring to FIG. 3, the thawed cells are centrifuged (step S32). That is, the thawed cells are transferred to a predetermined tube, then further mixed with the culture medium, and centrifuged. Thus, the supernatant is removed from the material produced as a result of the centrifugation. This centrifugation can be repeated one to several times. By removing the supernatant in this way, the upper thin layer can be stably removed while preserving the precipitate of the thawed cells.

[0062] Afterwards, referring to FIG. 3, a cell solution is prepared by adding a medium to the centrifuged cells (step S33). That is, the previously described IMDM medium is additionally provided and mixed with the centrifuged cells to prepare the cell solution. At this time, the mixing can be performed using, for example, a pipette. Meanwhile, the IMDM medium is additionally supplied to the cell solution mixed with the medium in this manner, and mixing is performed to finally prepare the cell solution.

[0063] Afterwards, referring to FIG. 3, the cell solution is provided to the SERS substrate array (step S34). That is, the finally prepared cell solution is provided to the SERS substrate array (401) prepared through the preceding step S20. At this time, the cell solution is provided sufficiently to allow the SERS substrate array (401) to be deposited. That is, as previously described, a stacked particle (310) in which nanoparticles are stacked is formed on each of the SERS substrates, and the cell solution is provided sufficiently to form a height greater than the height at which the stacked particle (310) is formed so as to allow the stacked particle (310) to be deposited.

[0064] Meanwhile, since a plurality of SERS substrates (400) are arranged in the SERS substrate array (401), the cell solution is provided to all of the SERS substrates (400) to allow the stacked particles (310) to be deposited. That is, with the SERS substrate array (401) formed to have a predetermined depth, the cell solution can be sufficiently provided to the entire SERS substrate array (401) so that all of the stacked particles (310) can be deposited.

[0065] Afterwards, referring to FIG. 3, culture of the cells is performed while replacing the medium (step S35). At this time, the timing of replacing the medium can be determined based on the change in the color of the medium, and the medium can be replaced repeatedly until sufficient culture of the cells is performed.

[0066] Thus, as illustrated in FIG. 4, the cell (500) is cultured in the same way on each SERS substrate (400) of the SERS substrate array (401).

[0067] Afterward, referring to FIG. 1, a drug is administered to the cultured cells (step S40). At this time, the administered drug performs a specific reaction on the rat brain endothelial cells as previously described, for example, the reaction may be a reaction that passes through the cell barrier. That is, as the drug is administered, the cultured cells may be induced to pass through.

[0068] At this time, the above drug must be provided to each SERS substrate (400) of the SERS substrate array (401). Meanwhile, since this embodiment tests the reactivity of the administered drug, the drugs administered to each SERS substrate (400) must ultimately be different drugs. Of course, to obtain more objective information regarding the same drug, the same drug may be administered to multiple SERS substrates.

[0069] Accordingly, if the number of types of drugs whose reactivity must be compared is equal to the number of SERS substrates (400), different drugs must be administered to each SERS substrate, but if the number of types of drugs is less than the number of SERS substrates, one drug may be administered to multiple SERS substrates.

[0070] Afterwards, referring to FIG. 1, Raman signals are measured from the cells to which the drug has been administered (step S50). Although not illustrated, these Raman signals are obtained through a Raman spectrometer. A more specific step for measuring the Raman signals (step S50) is described as follows.

[0071] FIG. 5 is a flowchart illustrating the step of measuring the Raman signal of FIG. 1. FIG. 6 is a graph to explain the step of optimizing the Raman signal measurement conditions of FIG. 5.

[0072] Referring to FIG. 5, in the step of measuring the Raman signal (step S50), first, the SERS substrate array (401) containing the cultured cells to which the drug has been administered is moved to a petri dish (step S51). At this time, the petri dish is already filled with saline solution, for example, phosphate-buffered saline (PBS) or DPBS (Dulbecco's phosphate-buffered saline).

[0073] After that, the SERS substrate array (401) transferred to the Petri dish is removed from the Petri dish, the saline solution is provided to the SERS substrate array (401), and the medium is removed, excluding the cultured cells to which the drug was administered (step S52).

[0074] At this time, the step of removing the medium (step S52) can be performed repeatedly from one to several times, thereby effectively removing all of the medium.

[0075] Afterward, the SERS substrate array (401) is moved to a measurement plate (step S53). In this case, only the cultured cells to which the drug has been administered are present on the SER substrate array (401), and all other substances, such as moisture, must be removed.

[0076] Afterward, the measurement conditions of the Raman signal using the Raman spectrometer are optimized (step S54). That is, when acquiring a Raman signal for a cultured cell to which a drug is administered, which is present in the SERS substrate array (401), using the Raman spectrometer, the acquired Raman signal may differ depending on the distance between the Raman spectrometer and the SERS substrate array (401). This is because the signal is affected as the distance to the measurement point reached by the laser provided through the Raman spectrometer varies.

[0077] Accordingly, in this embodiment, the measurement conditions of the Raman signal are optimized by varying the distance between the Raman spectrometer and the SERS substrate array (401). At this time, the optimization of the measurement conditions of the Raman signal means, as shown in FIG. 6, varying the distance between the SERS substrate array (401) and the Raman spectrometer to determine the distance at which the so-called R6G (Rhodamine 6G) signal is most clearly and significantly produced. Thus, the Raman spectrometer is positioned at that distance to optimize the measurement conditions.

[0078] In FIG. 6, the distance at which the R6G signal is most clearly and significantly emitted can be obtained as A, and the Raman spectrometer is adjusted to be located at that distance. Of course, as shown in A, two or more similar distances at which the R6G signal is most clearly and significantly emitted may occur, and the distance at which it is most significantly emitted is selected to determine the optimal measurement conditions at that distance.

[0079] Afterward, when the above measurement conditions are optimized, the Raman signal for the cultured cells administered with the drug is obtained using the Raman spectrometer (step S55).

[0080] That is, as previously explained, since cultured cells administered with different drugs exist on each SERS substrate (400) of the SERS substrate array (401), the reactivity of each drug to the cells can be confirmed by acquiring the Raman signal for each SERS substrate (400).

[0081] Thus, referring to FIG. 1, the optimal drug is selected based on the results of confirming the reactivity of each of the above drugs to the cells (step S60). At this time, the optimal drug may be the drug with the best penetration effect into the nerve cells when the cells are rat brain endothelial cells, as previously explained.

[0082] At this time, the step of measuring the Raman signal (step S50) may be performed repeatedly at predetermined intervals after the drug is administered. For example, the responsiveness of the drug to the cell following the administration of the drug may be determined in units of time or days. Thus, the measurement results of the Raman signal, which are repeatedly performed at predetermined intervals, may be accumulated to finally select the optimal drug (step S60).

[0083] Figures 7a to 7f are graphs showing the results of measuring Raman signals during the culture process of the cells under examination.

[0084] FIGS. 7a to 7f are graphs illustrating the results of measuring Raman signals using the Raman spectrometer according to the time of culture during the process of culturing the cells previously exemplified using rat brain endothelial cells.

[0085] For example, as shown in signal B in Fig. 7b, the growth state or change state of the cell during the culture process can be confirmed based on the occurrence of a new signal peak in the Raman signal when 2 hours have elapsed since the cell was cultured.

[0086] In addition, as shown in signal C in Fig. 7d, when 4 hours have elapsed since the cell was cultured, a new signal peak appears in the Raman signal, or the size of the existing peak (peak in Fig. 7b) rapidly increases or changes, and based on this, the growth state or change state during the cell culture process can be confirmed.

[0087] In addition, as shown in signal D in Fig. 7e, when 24 hours have elapsed since the cell was cultured, the growth state or change state during the cell culture process can be confirmed based on the occurrence of a new signal peak in the Raman signal or the increase or change in the size of existing peaks.

[0088] Likewise, as shown in signal E in Fig. 7f, when 60 hours have elapsed since the cell was cultured, the growth state or change state during the cell culture process can be confirmed based on the fact that the size of the existing peaks in the Raman signal increases or changes significantly.

[0089] As described above, as the brain blood vessel cells of the mouse are cultured, information regarding the degree of culture or culture characteristics of the brain blood vessel cells can be obtained based on the measurement results of Raman signals measured at preset time intervals.

[0090] In other words, if a drug that passes through the brain blood vessel cells is administered to the cultured cells, a drug passage test can be performed on the cultured brain blood vessel cells, and by acquiring Raman signals regarding the state at predetermined intervals, information regarding the characteristics of the cellular state of the brain blood vessel cells can be obtained.

[0091] Thus, as explained above, if the state of each brain blood vessel cell is periodically acquired via the Raman signal at preset intervals for the brain blood vessel cells to which various drugs have been administered simultaneously, information regarding the drug that most effectively passes through the brain blood vessel cells among the administered drugs can ultimately be obtained.

[0092] According to the embodiments of the present invention as described above, a SERS substrate array is fabricated to include a plurality of SERS substrates, allowing for simultaneous cell culture and the simultaneous determination of the drug effects of each drug by administering different drugs to each cultured cell, thereby improving the speed and relative comparative performance of drug responsiveness testing.

[0093] In particular, regarding the examination of the above-mentioned drug responsiveness, it was verified that the examination can be performed on mouse brain vascular cells based on Raman signals, and thus the functionality of the blood-brain barrier of mouse brain vascular cells can also be confirmed through Raman signals.

[0094] In addition, when fabricating the above SERS substrate, instead of growing nanoparticles directly on the substrate, nanoparticles are deposited on the substrate through sputtering to pass through the porous structure, thereby reducing the time required for nanoparticle deposition and enabling the fabrication of a more rapid and effective SERS substrate.

[0095] In addition, by optimizing the measurement conditions of the Raman signal before confirming the responsiveness of the cell by measuring the Raman signal, the accuracy of the signal can be further improved when confirming the responsiveness of the cell solely through the Raman signal.

[0096] Although the present invention has been described above with reference to preferred embodiments, those skilled in the art will understand that various modifications and changes can be made to the invention without departing from the spirit and scope of the invention as set forth in the following claims.

Claims

1. Step of fabricating the SERS substrate; A step of fabricating a SERS substrate array comprising a plurality of the above-mentioned SERS substrates; A step of culturing cells on the above-mentioned SERS substrate array; A step of administering a drug to the cultured cells; A step of measuring Raman signals from cells to which the above drug has been administered; and A drug responsiveness test method comprising the step of selecting the optimal drug among the administered drugs based on the measured Raman signal.

2. In Paragraph 1, The above cells are rat brain endothelial cells, and A drug responsiveness test method characterized in that the optimal drug is the drug with the best responsiveness to the cell.

3. In paragraph 2, in the step of selecting the optimal drug, A drug responsiveness test method characterized by determining that the responsiveness to the cell is excellent when a new peak is identified or the size of an existing peak changes in the measured Raman signal.

4. In paragraph 2, the reactivity to the cell refers to, A drug responsiveness testing method characterized by the property of passing through the cell barrier.

5. In the step of manufacturing the SERS substrate according to claim 1, A drug responsiveness testing method characterized by sputtering nanoparticles onto a base substrate and stacking the nanoparticles on the base substrate.

6. In Paragraph 5, A drug reactivity testing method characterized by the above nanoparticles passing through the pores of a porous substrate and sputtering onto the base substrate.

7. In Paragraph 6, The size of the above porous structure is larger than the nanoparticle, and A drug responsiveness test method characterized in that the arrangement of the above-mentioned holes is a random arrangement.

8. In the step of fabricating the SERS substrate array according to claim 1, A drug responsiveness testing method characterized by arranging the above SERS substrates in a predetermined number of columns and rows to fabricate the SERS substrate array on a single plate.

9. In claim 1, the step of culturing the cells is, Step of thawing frozen cells; A step of centrifuging the above-mentioned thawed cells; A step of preparing a cell solution by adding a culture medium to the centrifuged cells; The step of providing the cell solution to the SERS substrate array; and A drug responsiveness test method characterized by including the step of replacing the medium and culturing the cells.

10. In claim 9, in the step of centrifuging the thawed cells, A drug responsiveness test method characterized by adding a culture medium to the above-mentioned thawed cells and centrifuging to remove the supernatant.

11. In claim 9, at the step of providing the cell solution, A drug responsiveness testing method characterized by providing the above cell solution to precipitate the above SERS substrates.

12. In paragraph 1, the step of measuring the Raman signal is, A step of transferring the SERS substrate array containing cultured cells to which the above drug has been administered to a Petri dish; Step of removing the above-mentioned medium; A step of moving the above SERS substrate array to a measurement plate; A step of optimizing the Raman signal measurement conditions of a Raman spectrometer; and A drug responsiveness test method characterized by including the step of measuring the Raman signal under the above-described optimized measurement conditions.

13. In paragraph 12, the step of measuring the Raman signal is, A drug responsiveness test method characterized by performing the test repeatedly at predetermined intervals while the above-mentioned drug is administered.

14. In the step of optimizing the Raman signal measurement conditions of Clause 12, A drug responsiveness testing method characterized by varying the distance between the above-described Raman spectrometer and the above-described SERS substrate array.

15. In paragraph 14, the optimized Raman signal measurement conditions are, A drug responsiveness testing method characterized by the distance between the Raman spectrometer and the SERS substrate array at the case where the R6G (Rhodamine 6G) signal is most clearly and significantly emitted.

Images