A drug analysis platform based on lensless imaging technology

Abstract

The application discloses a drug analysis platform based on a lensless imaging technology, which combines the lensless imaging technology and microfluidic technology, can complete efficient and rapid cell imaging and activity analysis, realizes evaluation of dose-dependent cell reactions under various drug concentrations on a small area, and further completes low-error, low-cost and high-throughput drug screening work.

Description

Technical Field

[0001] This invention relates to a drug analysis platform based on lensless imaging technology, belonging to the field of microfluidic chip technology. Background Technology

[0002] Lensless microscopy refers to the recording of the projection of a sample above an imaging chip using an image sensor such as a CCD or CMOS sensor located beneath the chip, without the need for optical lenses. When cells are cultured directly attached to the imaging area of ​​the lensless imaging chip, the light source directly illuminating the cells above the imaging area produces the highest quality cell projection. The image sensor directly below the imaging area records the cell projection, and the image information is transmitted through surrounding electronic components and circuits. Digital imaging techniques and mathematical calculations are then used to reconstruct the cell projection, ultimately yielding a clear cell image. Although the imaging area is small, this technology can clearly capture images of tens of thousands of cells within that area, achieving high-resolution, wide-field-of-view microscopic imaging of live cells. Traditional live-cell imaging techniques primarily use optical microscopes. Unlike direct projection and digital imaging, these rely on lenses to refract and focus light from an object onto an imaging plane to form an image. This approach cannot simultaneously achieve high resolution and a wide field of view, making it difficult to monitor large-scale dynamic responses of cells, quantify cell viability, or capture dynamic changes such as cell growth and migration. Lensless microscopy overcomes the disadvantages of traditional optical microscopes in live cell imaging through its unique imaging technology. In the field of drug analysis, it analyzes cell viability by observing the growth and migration of live cells and statistically analyzing the optical morphology and confluence of cells, thereby evaluating cell drug response.

[0003] Microfluidic chip technology, as an analytical technique capable of precisely controlling fluid flow, boasts advantages such as miniaturization, high efficiency, speed, and low cost, and is widely used in the field of drug analysis. Among these, concentration gradient microfluidic chips are a crucial technique in drug analysis. This technology constructs drug concentration gradients to study dose-dependent cellular responses at different drug concentrations, enabling high-throughput drug screening. Traditional drug concentration gradient assays are primarily performed on multi-well plates, but this method typically requires manual pipetting, resulting in high workload, susceptibility to errors, and high reagent consumption. Concentration gradient microfluidic chips, on the other hand, allow the introduction of different drug concentrations at different inlets. These drug fluids are then automatically split and mixed layer by layer, forming a precise and stable concentration gradient at the outlet. Therefore, concentration gradient microfluidic chips are widely used for constructing drug concentration gradients due to their advantages such as automated pipetting, low error, and low reagent consumption.

[0004] Currently, most concentration gradient microfluidic chips are combined with traditional live-cell imaging techniques to assess dose-dependent cellular responses by monitoring cell viability and to screen potential drugs and optimal drug concentrations based on the assessment results. However, due to the limitations of traditional optical microscopy, this method often requires a large imaging area to provide sufficient cell samples for drug analysis, resulting in complex experimental conditions, difficult data analysis, and hindering efficient drug screening. Therefore, combining concentration gradient microfluidic chips with lensless microscopy technology promises to complete the screening of several drugs in a smaller imaging area compared to traditional drug analysis platforms, greatly improving drug screening efficiency. However, the size of concentration gradient microfluidic chips is often larger than that of lensless imaging chips, making it difficult to match the packaging area of ​​lensless imaging chips. Currently, commonly used concentration gradient microfluidic chips often adopt a "Christmas tree" structure. Figure 1 This structure, shaped like a Christmas tree, consists of multiple main channels from top to bottom, with each layer connected by multiple branch channels. Drugs of different concentrations are injected from the top layer and then distributed to the branches within each main channel, mixing to create new concentrations and ultimately forming a concentration gradient at the bottom. The tortuous structure of the branch channels facilitates thorough drug mixing. However, this structure suffers from drawbacks such as large area requirements, the channels being concentrated on one side, numerous tortuous channels, and difficulties in channel optimization, making integration with lensless imaging chips challenging. Furthermore, because the imaging area of ​​a lensless imaging chip functions differently from its surrounding area, their materials and surface structures often differ, making close integration with concentration gradient microfluidic chips difficult. Therefore, combining concentration gradient microfluidic chips with lensless microscopy to overcome the technical challenges of drug analysis platforms built on traditional live-cell imaging techniques has become a pressing issue in this field. Summary of the Invention

[0005] Objectives of the Invention: The first objective of this invention is to provide a concentration gradient microfluidic chip. The second objective of this invention is to provide a drug analysis platform suitable for the aforementioned concentration gradient microfluidic chip without lens imaging technology.

[0006] Technical Solution: This invention provides a concentration gradient microfluidic chip, which is formed by bonding a flow layer and a connecting layer. The flow layer has a first inlet and a second inlet at its two ends, and a groove on its lower surface. After the connecting layer and the flow layer are bonded, a fluid channel is formed. The fluid channel is connected to a through hole in the center of the connecting layer and is disposed around the through hole. There are at least two through holes. The fluid channel includes different branch channels connected in parallel and / or in series, so that the drug solution or diluent forms a concentration gradient when it reaches the outlet corresponding to the different through holes from the first inlet or the second inlet.

[0007] Furthermore, the branch channel comprises flow channels of different lengths, and the flow channels of different lengths are achieved by different bending lengths and bending times.

[0008] Furthermore, the design formula for the fluid channel is as follows:

[0009]

[0010] In the formula, Q represents the flow rate, with units of m³. 3 / s; Δp is the pressure drop, in Pa; R F Flow resistance, unit is Pa·s / m 3 ;

[0011]

[0012] In the formula, Δp n R represents the pressure drop across each fluid channel, measured in Pa. Fn The flow resistance of each channel is expressed in Pa·s / m. 3 Q n The flow rate of each fluid channel is expressed in m³. 3 / s, where m is the number of outlets corresponding to the through holes;

[0013]

[0014] In the formula, L n The length of each flow channel is in meters (m).

[0015] Furthermore, on one side of the first or second injection port, two branch channels branch out through the fluid channel and connect to through hole 1 and through hole 2 respectively; on the other side of the first or second injection port, 1, 2, 3...m-2 branch channels branch out and connect to through holes 3, 4, 5...m-2 respectively, where m is the number of through holes.

[0016] Furthermore, assuming the diluent enters from the first inlet and the drug enters from the second inlet, the diluent flows through L1+L4 to reach outlet 1 corresponding to the through hole; the drug flows through L7+L6 to reach outlet m corresponding to the through hole.

[0017] When the number of outlets corresponding to the through-holes is m=4, the diluent flows through L8 and through L7+L A3 The medicinal liquids merge and then flow through L B3 +L E3 Then it reaches the outlet 3 corresponding to the through hole; the diluent flows through L8 and through L7+L A3 The medicinal liquids merge and then flow through L B3 +L5 then merges with the diluent flowing through L1+L2, and then flows through L3 to reach the outlet 2 corresponding to the through hole;

[0018] When the number of outlets corresponding to the through-holes is m=5, the diluent flows through L8 and through L7+L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L D3 +L E3 Then it reaches the outlet 3 corresponding to the through hole; the diluent flows through L8 and through L7+L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L D3 After L5, it merges with the diluent flowing through L1+L2, then flows through L3 to reach the outlet 2 corresponding to the through hole; the diluent flows through L8 and the diluent flowing through L7+L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L C3 With flow through L7+L A4 +L B4 The medicinal liquids merge and then flow through L E4 Reach the outlet 4 corresponding to the through hole;

[0019] When the number of outlets corresponding to the through-holes is m=6, the diluent flows through L8 and through L7+L A5 +L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L D3 +L E3 Then it reaches the outlet 3 corresponding to the through hole; the diluent flows through L8 and through L7+L A5 +L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L D3 After L5, it merges with the diluent flowing through L1+L2, then flows through L3 to reach the outlet 2 corresponding to the through hole; the diluent flows through L8 and the diluent flowing through L7+L A5 +L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L C3 With flow through L7+L A5 +L A4 +L B4 The medicinal liquids merge and then flow through L D4 +L E4 Reaching outlet 4 corresponding to the through hole; the diluent flows through L8 and through L7+L A5 +L A4 +L A3 The medicinal liquids merge and then flow through L B3 +L C3 With flow through L7+L A5 +LA4 +L B4 The medicinal liquids merge and then flow through L D4 +L C4 With flow through L7+L A5 +L B5 The medicinal liquids merge and then flow through L E5 Then it reaches the outlet 5 corresponding to the through hole;

[0020] When the export quantity m > 6, the same logic applies;

[0021] The relationship between the lengths of each flow channel is shown below:

[0022] Q2L2 + Q3L3 = Q4L4

[0023] Q5L5 + Q3L3 = Q E3 L E3

[0024]

[0025] Q E3 L E3 +Q D3 L D3 +Q B3 L B3 +Q8L8=L1+L4

[0026]

[0027] Q A3 L A3 +Q C3 L C3 +Q B3 L B3 =Q B4 L B4

[0028] Q E3 L E3 +Q D3 L D3 =Q C3 L C3 +Q D4 L D4 +Q E4 L E4

[0029] Q E(m-2) L E(m-2) =Q C(m-2) L C(m-2) +Q E(m-1) L E(m-1)

[0030] Q An L An +QBn L Bn +Q Cn L Cn +Q Dn L Dn =Q B(n+1) L B(n+1) (n = 4, ..., m - 2)

[0031] Q En L En =Q Cn L Cn +Q D(n+1) L D(n+1) +Q E(n+1) L E(n+1) (n = 4, ..., m-3)

[0032]

[0033] In the formula, s is the exit number, s = 3, 4, 5, ..., m-1.

[0034] Furthermore, the groove has a width of 100μm and a height of 100μm.

[0035] Furthermore, the connecting layer and the flow layer are of equal size.

[0036] Furthermore, the number of through holes is 4 to 9.

[0037] The present invention also provides a drug analysis platform based on lensless imaging technology. The drug analysis platform includes a lensless imaging chip, a cell chamber layer, and the above-mentioned concentration gradient microfluidic chip, which are bonded and assembled in a bottom-to-top order. The lensless imaging chip has a lensless imaging region. The cell chamber layer has through holes corresponding to the positions of the connecting layers. After bonding, a cell culture chamber is formed, and the position of the cell culture chamber is within the lensless imaging region.

[0038] Furthermore, the cell chamber layer and the lensless imaging region have the same area and are smaller than the size of the concentration gradient microfluidic chip.

[0039] The area of ​​the cell chamber layer should be designed to be slightly larger than the lensless imaging area. If the area of ​​the cell chamber layer is smaller than the lensless imaging area, the imaging area cannot be maximized. If the area of ​​the cell chamber layer is too large, it may not fit properly due to the unevenness of the lensless imaging area.

[0040] Furthermore, the dimensions of the cell chamber layer are 3-10 mm in length, 3-10 mm in width, and ≥1 mm in height; the dimensions of the flow layer and the connecting layer are 29-37 mm in length, 23-27 mm in width, and ≥1 mm in height; and the diameter of the through holes on the connecting layer and the cell chamber layer is 1-3 mm.

[0041] Furthermore, the height of the cell chamber layer is 1–5 mm; the height of the flow layer and the connecting layer is 1–4 mm.

[0042] Furthermore, the cell chamber layer, the connecting layer, and the flow layer are all made of polydimethylsiloxane.

[0043] Furthermore, the cell culture chamber is used to establish neuronal cell lines, primary neurons, and glial cell models.

[0044] Beneficial Effects: Compared with existing technologies, this invention has the following significant advantages: This invention provides a compact and lightweight drug analysis platform based on lensless imaging technology. By designing a connecting layer and a cell chamber layer, the flow layer and the lensless imaging chip are indirectly connected, solving the difficulty that the concentration gradient microfluidic chip cannot be directly and tightly attached to the lensless imaging chip due to the different materials and surface structures of the lensless imaging area and its surrounding chip area. This achieves the combination of concentration gradient microfluidic chip and lensless imaging technology. An annular fluid channel is designed around the central area on the flow layer corresponding to the bottom lensless imaging area. This ensures that the fluid channel does not obstruct the lensless imaging area, making the fluid channel uniformly distributed while minimizing the flow channel area. This saves space in the outer packaging area of ​​the lensless imaging chip, allowing for changes in the flow channel according to drug analysis needs. The length of each flow channel is accurately calculated using the equivalent circuit method, enabling the concentration gradient microfluidic chip to generate a stable concentration gradient. The through-hole in the central area of ​​the platform allows the light source above the platform to be unobstructed while culturing cells, directly illuminating the attached cells on the lensless imaging chip to complete projection imaging. This platform can simultaneously monitor cell growth and migration activities while applying different concentrations of drug to cells in each chamber, thereby analyzing cell viability to assess dose-dependent cellular responses. The platform integrates a concentration gradient microfluidic chip and a lensless imaging chip, and through the design of an array of through-holes, it maximizes the imaging area of ​​the lensless imaging chip. This allows the lensless imaging chip, which could originally only hold one concentration of drug, to simultaneously analyze multiple concentrations of drug, greatly improving drug screening efficiency and enabling low-cost, low-error, high-throughput drug screening. Attached Figure Description

[0045] Figure 1 This is a diagram combining a traditional concentration gradient microfluidic chip with a lensless imaging chip;

[0046] Figure 2 This is a physical image of a drug analysis platform based on lensless imaging technology;

[0047] Figure 3This is a front view of a drug analysis platform based on lensless imaging technology;

[0048] Figure 4 This is a top view of a drug analysis platform based on lensless imaging technology;

[0049] Figure 5 This is a top view of the lensless imaging chip of this drug analysis platform based on lensless imaging technology;

[0050] Figure 6 This is a schematic diagram of the fluid channel structure of a drug analysis platform based on lensless imaging technology;

[0051] Figure 7 This is a schematic diagram of the four-chamber flow layer of a drug analysis platform based on lensless imaging technology;

[0052] Figure 8 This is a schematic diagram of the five-chamber flow layer of this drug analysis platform based on lensless imaging technology;

[0053] Figure 9 This is a schematic diagram of the connection layer of a drug analysis platform based on lensless imaging technology;

[0054] Figure 10 This is a schematic diagram of the cell chamber layer of a drug analysis platform based on lensless imaging technology;

[0055] Figure 11 This is a simulation diagram of the concentration gradient in the four chambers of a drug analysis platform based on lensless imaging technology;

[0056] Figure 12 This is a simulation diagram of the concentration gradient in the five chambers of a drug analysis platform based on lensless imaging technology;

[0057] Figure 13 This is a fluorescent dye effect diagram of the fluid channel of a drug analysis platform based on lensless imaging technology;

[0058] Figure 14 This is a fluorescent dye spectrum of the fluid channel of a drug analysis platform based on lensless imaging technology;

[0059] Figure 15 The images show cell cultures in circular wells of the same size as the cell culture chambers of this lensless imaging-based drug analysis platform. The top image shows HT22 cells, and the bottom image shows BV2 cells. Detailed Implementation

[0060] The technical solution of the present invention will be further described below with reference to the accompanying drawings.

[0061] Example 1

[0062] The structure of the drug analysis platform in this embodiment is referenced. Figures 2 to 10 . Figure 2 The image shown is a physical illustration of the drug analysis platform of the present invention. Taking a four-cell culture chamber and a five-cell culture chamber as examples, the drug platform is assembled from a lensless imaging chip 1, a cell chamber layer 3, a connecting layer 4, and a flow layer 5 in a bottom-to-top order, after aligning their geometric centers and bonding them together. The cell chamber layer 3, connecting layer 4, and flow layer 5 are all made of polydimethylsiloxane with light-transmitting properties. The lensless imaging region 2 is the same size as the cell chamber layer 3; the connecting layer 4 is the same size as the flow layer 5, and larger than the size of the lensless imaging region 2.

[0063] The lensless imaging chip 1 has a lensless imaging area 2.

[0064] A concentration gradient microfluidic chip is formed by bonding the connecting layer 4 and the flow layer 5. The flow layer 5 has a first inlet and a second inlet at its two ends, respectively. A groove is formed on the lower surface of the flow layer 5, creating a fluid channel after the bonding of the connecting layer 4 and the flow layer 5. The groove has a width of 100 μm and a height of 100 μm. A circular through-hole is provided on the connecting layer 4. The fluid channel is connected to the through-hole in the center of the connecting layer and is positioned around the through-hole. The fluid channel includes different branch channels connected in parallel and / or in series, so that the drug solution or diluent presents a concentration gradient when it reaches different through-holes from the first or second inlet. Both the connecting layer 4 and the flow layer 5 have dimensions of 29 mm in length, 23 mm in width, and 4 mm in height.

[0065] The cell chamber layer 3 has dimensions of 10 mm in length, 10 mm in width, and 5 mm in height. The cell chamber layer 3 has through-holes corresponding to the positions of the connecting layer 4, which, when bonded together, form the cell culture chamber 6. The cell culture chamber 6 has a diameter of 3 mm.

[0066] The concentration gradient microfluidic chip of the present invention combines the characteristics of lensless imaging chip. It designs an annular flow channel by means of equivalent circuit method, and sets the lensless imaging area in the center of the flow channel area. Under the condition that the solution at each outlet forms a concentration gradient and the flow rate is the same, it achieves the effect of uniform flow channel arrangement and compact structure.

[0067] The equivalent circuit method refers to equating the pressure drop, flow rate, and flow resistance of each flow channel to the voltage, current, and resistance in a closed circuit.

[0068] The pressure drop in the flow channel is related to the flow rate and flow resistance as follows:

[0069]

[0070] In the formula, Q represents the flow rate, with units of m³. 3 / s; Δp is the pressure drop, in Pa; R F Flow resistance, unit is Pa·s / m3 .

[0071] According to Kirchhoff's laws, the algebraic sum of the currents at each node in a closed circuit is 0, and the algebraic sum of the voltages in each loop is 0. Therefore:

[0072]

[0073] In the formula, Δp n R represents the pressure drop across each flow channel, expressed in Pa. Fn The flow resistance of each channel is expressed in Pa·s / m. 3 Q n The flow rate of each channel is expressed in m³. 3 / s.

[0074] When the width and height of the flow channel are uniform, the flow resistance is directly proportional to the length of the flow channel. Therefore:

[0075]

[0076] In the formula, L n The length of each flow channel is in meters (m).

[0077] During drug injection, it is necessary to ensure synchronous injection into each cell culture chamber to guarantee the same injection volume and generate the required linear concentration gradient, thereby achieving different drug concentrations in each chamber and realizing high-throughput drug screening. Therefore, assuming the initial drug flow rates at the two injection ports are the same, the initial drug flow rate to be allocated to each channel is calculated based on the required results (including the injection rate and concentration in each chamber). This ensures that the two initial drug concentrations are mixed at each node according to the preset flow rate ratio, ultimately forming a concentration gradient. Furthermore, the height and width of each channel are controlled to be consistent, and the length of each channel is calculated based on the flow rate and Kirchhoff's laws to adjust the flow resistance and control the flow rate.

[0078] like Figure 6 As shown, when the number of cell culture chambers is m, the relationship between the lengths of each channel is as follows:

[0079] Q2L2 + Q3L3 = Q4L4

[0080] Q5L5 + Q3L3 = Q E3 L E3

[0081]

[0082] Q E3 L E3 +Q D3 L D3 +Q B3 L B3+Q8L8=L1+L4

[0083]

[0084] Q A3 L A3 +Q C3 L C3 +Q B3 L B3 =Q B4 L B4

[0085] Q E3 L E3 +Q D3 L D3 =Q C3 L C3 +Q D4 L D4 +Q E4 L E4

[0086] Q E(m-2) L E(m-2) =Q C(m-2) L C(m-2) +Q E(m-1) L E(m-1)

[0087] Q An L An +Q Bn L Bn +Q Cn L Cn +Q Dn L Dn =Q B(n+1) L B(n+1) (n = 4, ..., m - 2)

[0088] Q En L En =Q Cn L Cn +Q D(n+1) L D(n+1) +Q E(n+1) L E(n+1) (n = 4, ..., m-3)

[0089]

[0090] The following are the specific parameters of the flow layer in the four-cell and five-cell culture chambers. The fluid channel is designed within a rectangular area with a length of 24 mm and a width of 19 mm. The solution concentrations at the upper and lower inlets are 0 and 1 mol / m³, respectively. 3 .

[0091] See Figure 7 The length relationships of the flow channels in the four-chamber flow layer are shown below:

[0092]

[0093] The lengths of each channel in this four-chamber flow layer example are shown in the table below:

[0094] flow channel <![CDATA[L1]]> <![CDATA[L2]]> <![CDATA[L3]]> <![CDATA[L4]]> <![CDATA[L5]]> <![CDATA[L6]]> <![CDATA[L7]]> <![CDATA[L8]]> <![CDATA[L A3 ]]> <![CDATA[L B3 ]]> <![CDATA[L E3 ]]> Length / mm 23.3 3.9 10.5 12 26 43.5 17 17 5 10 23.5

[0095] See Figure 8 The length relationships of the channels in the five-chamber flow layer are shown below:

[0096]

[0097] The lengths of each channel in this five-chamber flow layer example are shown in the table below:

[0098]

[0099] Example 2

[0100] To evaluate the effectiveness of the fluid channel 8 constructed using the equivalent circuit method, the flow layer 5 was modeled using AutoCAD 2023 software, and a simulation experiment was conducted using COMSOL Multiphysics 6.0 software at a flow rate of 50 μL / min. The solution concentrations at the upper and lower inlets were 0 and 1 mol / m³, respectively. 3 See also Figure 11 and Figure 12 The results showed that the solution flow rates in the four-cell culture chamber and the five-cell culture chamber were similar, allowing for synchronous sample injection. Furthermore, the concentrations in the four-cell culture chamber were 0 and 0.33 mol / m³, respectively. 3 0.67mol / m 3 1mol / m 3 The concentrations in the five-cell culture chambers were 0 and 0.23 mol / m³, respectively. 3 0.51 mol / m 3 0.76mol / m 3 1mol / m 3 All of them form a stable linear concentration gradient.

[0101] Specifically, in the four-cell culture chamber, a solution with a concentration of 0 enters through the upper inlet (first inlet), flows through L1+L4, and reaches outlet 1; a solution with a concentration of 0 enters through the upper inlet, flows through L8, and then merges with a solution entering through the lower inlet (second inlet) flowing through L7+L4. A3 The concentration is 1 mol / m 3 The solutions merge, and then pass through L B3 +L E3The solution reaches outlet 3; a solution with a concentration of 0 enters from the upper inlet, flows through L8, and then flows through L7+L from the lower inlet (second inlet). A3 The concentration is 1 mol / m 3 The solutions merged, and after L B3 Then it passes through L5 and merges with the solution with a concentration of 0 that has passed through L1+L2, and then passes through L3 to reach outlet 2; the concentration is 1 mol / m³. 3 The solution enters from the lower inlet, flows through L7+L6 and reaches outlet 4.

[0102] In the five-cell culture chamber, a solution with a concentration of 0 enters through the upper inlet (first inlet), flows through L1+L4, and reaches outlet 1; a solution with a concentration of 0 enters through the upper inlet, flows through L8, and enters through the lower inlet (second inlet), flowing through L7+L4. A4 +L A3 The concentration is 1 mol / m 3 The solutions merge and flow through L B3 +L D3 +L E3 The solution then enters through outlet 3; a solution with a concentration of 0 enters through the upper inlet and flows through L8, while a solution entering through the lower inlet (second inlet) flows through L7+L. A4 +L A3 The concentration is 1 mol / m 3 The solutions merge and flow through L B3 +L D3 After L5, it merges with the 0-concentration solution flowing through L1+L2 from the upper inlet (first inlet), and then flows through L3 into outlet 2; the 0-concentration solution enters from the upper inlet and flows through L8, and merges with the solution entering from the lower inlet (second inlet) and flowing through L7+L... A4 +L A3 The concentration is 1 mol / m 3 The solutions merge and flow through L B3 +L C3 Then, the flow entering from the lower injection port (second injection port) passes through L7+L. A4 +L B4 The concentration is 1 mol / m 3 The solutions merge and then flow through L E4 Enter through outlet 4; enter through the lower injection port (second injection port) with a concentration of 1 mol / m 3 The solution flows through L7+L6 to reach outlet 5.

[0103] Example 3

[0104] The formation of the concentration gradient was further verified by measuring the UV-Vis absorption spectrum of Rhodamine 6G. First, a 0.15 mM Rhodamine 6G solution was prepared, and then it and deionized water were injected separately through different inlets using a syringe pump. The results were as follows: Figure 12 As shown (taking a four-cell culture chamber as an example), the absorbance of the solution in each cell culture chamber 6 was measured. The absorbance of the four cell culture chambers 6 at a wavelength of 527 nm is shown below. Figure 14 As shown, the results indicate that the absorbance of the solution in each chamber exhibits a linear relationship, consistent with the simulated experimental results.

[0105] Example 4

[0106] To demonstrate that the 3mm diameter cell culture chamber 6 can meet the requirements for cell culture, HT22 cells and BV2 cells, commonly used for drug screening in neurodegenerative diseases, were cultured for 24 hours in 3mm diameter round wells with a polydimethylsiloxane substrate, at a cell density of 1×10⁶ cells / mL. 5 The results showed that both cell types could grow normally in the round wells.

[0107] Example 5

[0108] A four-chamber drug analysis platform based on lensless imaging technology, with each chamber having a cell imaging area of ​​0.07065 cm². 2 While maintaining a resolution comparable to 400 times that of traditional optical microscopy, the field of view for live-cell imaging is increased by 35.325 times compared to traditional drug analysis platforms. By integrating concentration gradient microfluidic chip technology with lensless imaging technology, high-throughput cell imaging is achieved, reducing errors in quantifying cell viability and improving the efficiency of drug screening.

Claims

1. A drug analysis platform based on lensless imaging technology, characterized in that, The drug analysis platform comprises a lensless imaging chip, a cell chamber layer, and a concentration gradient microfluidic chip, which are bonded and assembled in a bottom-to-top order. The lensless imaging chip has a lensless imaging region. The cell chamber layer has through holes corresponding to the positions of the connecting layers. After bonding, a cell culture chamber is formed, and the position of the cell culture chamber is within the lensless imaging region. The concentration gradient microfluidic chip is composed of a flow layer and a connecting layer bonded together. The flow layer has a first inlet and a second inlet at its two ends, and a groove on its lower surface. After the connecting layer and the flow layer are bonded together, a fluid channel is formed. The fluid channel is connected to a through-hole in the center of the connecting layer and is located around the through-hole. There are at least two through-holes. The fluid channel includes different branch channels connected in parallel or / and in series, so that the drug solution or diluent presents a concentration gradient when it reaches the outlets corresponding to different through-holes from the first inlet or the second inlet. The design formula for the fluid channel is as follows: ； In the formula, Q represents the flow rate, with units of m³. 3 / s; Δp is the pressure drop, in Pa; R F Flow resistance, unit is Pa·s / m 3 ; ； In the formula, Δp n R represents the pressure drop across each flow channel, expressed in Pa. Fn The flow resistance of each fluid channel is expressed in Pa·s / m. 3 Q n The flow rate of each fluid channel is expressed in m³. 3 / s, where m is the number of outlets corresponding to the through holes; ； In the formula, L n The length of each flow channel is in meters (m).

2. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, The branch channel consists of flow channels of different lengths, and the different lengths of flow channels are achieved by different bending lengths and bending times.

3. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, Two branch channels branch out from one side of the first or second injection port through the fluid channel, respectively connecting to through hole 1 and through hole 2; 1, 2, 3...m-2 branch channels branch out from the other side of the first or second injection port, respectively connecting to through holes 3, 4, 5...m-2, where m is the number of through holes.

4. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, The connecting layer and the flow layer are of equal size.

5. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, The cell chamber layer and the lensless imaging region have the same area and are smaller than the size of the concentration gradient microfluidic chip.

6. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, The dimensions of the cell chamber layer are 3-10 mm in length, 3-10 mm in width, and ≥1 mm in height; the dimensions of the flow layer and the connecting layer are 29-37 mm in length, 23-27 mm in width, and ≥1 mm in height; the diameter of the through holes on the connecting layer and the cell chamber layer is 1-3 mm.

7. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, The cell chamber layer, connecting layer, and flow layer are all made of polydimethylsiloxane.

8. The drug analysis platform based on lensless imaging technology according to claim 1, characterized in that, The cell culture chamber is used to establish neuronal cell lines, primary neurons, and glial cell models.

Images