Microfluidic sorting chip, droplet screening method, system and equipment and storage medium
Patent Information
- Application Number
- CN202280102775.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2022-12-30
- Publication Date
- 2025-07-29
AI Technical Summary
Existing technology cannot provide diversified enzyme screening solutions and is difficult to meet the diverse requirements for enzyme performance in different usage scenarios.
By collecting a variety of electrical signals from the droplets to be tested on the microfluidic sorting chip, the target electrical signal that meets the intensity threshold is determined, and the characteristics of the droplets are judged based on the signal width and characteristic ratio, providing a variety of screening solutions.
It improves the flexibility and accuracy of enzyme screening and provides diversified screening solutions, suitable for scenarios such as enzyme screening, cell line breeding, and monoclonal antibody development.
Smart Images
Figure CN120390871A_ABST
Abstract
Description
Microfluidic sorting chip, droplet screening method, system, equipment and storage medium Technical Field
[0001] The present disclosure relates to the field of microfluidics technology, for example, to a microfluidics sorting chip, a droplet screening method, a system, a device and a storage medium. Background Art
[0002] Enzymes are a type of macromolecular biocatalyst with important applications in various fields, including industry and medicine. Different usage scenarios require different enzyme performance, such as thermal stability, stability in organic solvents, catalytic efficiency, substrate specificity, and so on. Therefore, it is very important to screen for enzymes with good results. In the enzyme screening process based on microdroplet technology, single cells and reaction substrates are encapsulated in microdroplets for reaction. Among them, the key step is to amplify, collect and process the signals of the droplets in specific bands, and make decisions and sorting based on the characteristics of each signal. At the same time, the enzyme screening method based on microdroplet technology can be applied to scenarios such as cell line breeding and monoclonal antibody development.
[0003] Existing technical means include enzyme labeling screening technology, spectrophotometer screening technology, gel electrophoresis screening technology, flow cytometer screening technology and microfluidic single-channel screening technology, etc. The above technical means have the defect of being unable to provide diversified screening solutions.
[0004] Summary of the Invention
[0005] The technical problem to be solved by the present disclosure is to overcome the defect that the existing technology cannot provide diversified screening solutions, and to provide a microfluidic sorting chip, droplet screening method, system, equipment and storage medium.
[0006] The present disclosure solves the above technical problems through the following technical solutions:
[0007] In a first aspect, the present disclosure provides a droplet screening method, the method comprising:
[0008] collecting at least two electrical signals of the droplet to be tested at a preset frequency, wherein each electrical signal of the droplet to be tested is obtained in a different analog-to-digital conversion signal channel;
[0009] determining a target electrical signal in the electrical signals that meets a strength threshold;
[0010] When the signal width of the target electrical signal meets the signal width threshold, extracting features from the target electrical signal data, determining whether the intra-channel feature of the target electrical signal meets a first preset condition and determining whether the inter-channel feature of the target electrical signal meets a second preset condition;
[0011] If yes, the droplet to be tested is determined to be a target droplet;
[0012] The intra-channel characteristics are characteristics of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels, and the inter-channel characteristics are characteristics of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels.
[0013] Optionally, the signal width is obtained by calculating the number of target electrical signals that meet an intensity threshold and an acquisition frequency.
[0014] Optionally, the step of determining whether the intra-channel feature of the target electrical signal satisfies a first preset condition includes:
[0015] Obtaining an average intensity value and a peak intensity value of the target electrical signal that meets a signal width threshold;
[0016] When the in-channel feature of the target electrical signal that meets the signal width threshold meets the first ratio threshold, the first preset condition is met;
[0017] The intra-channel feature is the ratio of the average intensity to the peak intensity of the target electrical signal that meets the signal width threshold.
[0018] Optionally, the inter-channel feature includes a first inter-channel feature and a second inter-channel feature, and the step of determining whether the inter-channel feature of the target electrical signal meets a second preset condition includes:
[0019] Acquire the target electrical signal that meets the first preset condition and obtain the intensity peak;
[0020] When the first inter-channel feature of the target electrical signal that meets the first preset condition meets the second ratio threshold, the second preset condition is met;
[0021] The inter-channel characteristic is a ratio of the peak values of the target electrical signals corresponding to the analog-to-digital conversion signal channels that meet a first preset condition.
[0022] and / or,
[0023] Acquire the target electrical signal that meets the first preset condition and obtain an average strength value;
[0024] When the second inter-channel feature of the target electrical signal that meets the first preset condition meets the third ratio threshold, the second preset condition is met;
[0025] The inter-channel characteristic is a ratio of the average intensity of the target electrical signal corresponding to each of the analog-to-digital conversion signal channels that meets a first preset condition.
[0026] Optionally, the step of collecting at least two electrical signals of the droplet to be tested at a preset frequency includes:
[0027] Acquire an optical signal of the droplet to be measured by a photomultiplier tube, collect the optical signal at a preset frequency and acquire at least two electrical signals of the droplet to be measured;
[0028] The optical signal includes at least one of a scattered light signal, a fluorescent signal and an absorbed light signal.
[0029] Optionally, the step of determining a target electrical signal in the electrical signal that meets a strength threshold further includes:
[0030] When at least two electrical signals of the droplet to be tested collected at a preset frequency meet an intensity threshold for the first time, the electrical signals are collected into a data packet;
[0031] until the electrical signal is lower than a strength threshold;
[0032] The data packet collects all electrical signals of at least two electrical signals of the droplet to be tested from the first time the signal meets the intensity threshold to the last time the signal meets the intensity threshold according to a preset frequency.
[0033] In the second aspect, the present disclosure provides a microfluidic sorting chip, which includes a sorting component and at least two branch channels. When the droplet to be tested is determined to be a target droplet by any of the droplet screening methods described above, the sorting component is used to receive a deflection instruction and sort the droplet to be tested into the corresponding branch channel.
[0034] In a third aspect, the present disclosure provides a droplet screening system, comprising:
[0035] an acquisition module, configured to acquire at least two electrical signals of the droplet to be tested at a preset frequency, wherein each electrical signal of the droplet to be tested is acquired in a different analog-to-digital conversion signal channel;
[0036] a determination module, configured to determine a target electrical signal in the electrical signal that meets a strength threshold;
[0037] a judgment module, configured to extract features from the target electrical signal data when the signal width of the target electrical signal meets a signal width threshold, judge whether the intra-channel feature of the target electrical signal meets a first preset condition, and judge whether the inter-channel feature of the target electrical signal meets a second preset condition;
[0038] If yes, the droplet to be tested is determined to be a target droplet;
[0039] The intra-channel characteristics are characteristics of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels, and the inter-channel characteristics are characteristics of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels.
[0040] In a fourth aspect, the present disclosure provides an electronic device comprising a memory, a processor, and a computer program stored in the memory and configured to run on the processor, wherein the processor implements any of the above-described droplet screening methods when executing the computer program.
[0041] In a fifth aspect, the present disclosure provides a computer-readable storage medium having a computer program stored thereon, wherein the computer program, when executed by a processor, implements any of the above-described droplet screening methods.
[0042] The positive progress of the present disclosure lies in: by collecting at least two electrical signals of a droplet to be tested at a preset frequency and determining a target electrical signal in the electrical signals that meets an intensity threshold, when the signal width of the target electrical signal meets the signal width threshold, extracting features from the target electrical signal data, and determining whether the droplet to be tested is a target droplet by judging whether the intra-channel feature of the target electrical signal meets a first preset condition and whether the inter-channel feature of the target electrical signal meets a second preset condition. This provides a variety of screening options for droplets to be tested, improving the flexibility and accuracy of screening droplets to be tested. BRIEF DESCRIPTION OF THE DRAWINGS
[0043] FIG1 is a flow chart of a droplet screening method provided in Example 1 of the present disclosure.
[0044] FIG2 is a schematic diagram of a structure for performing droplet screening using a microfluidic sorting chip as provided in Example 1 of the present disclosure.
[0045] FIG3 is a partial flow chart of a droplet screening method provided in Example 1 of the present disclosure.
[0046] FIG4 is a partial flow chart of another droplet screening method provided in Example 1 of the present disclosure.
[0047] FIG5 is a partial flow chart of another droplet screening method provided in Example 1 of the present disclosure.
[0048] FIG6 is a flowchart of droplet screening using two signals from channel 1 and channel 3, provided in Example 1 of the present disclosure.
[0049] FIG7 is a schematic diagram of a module of a droplet screening system provided by an embodiment of the present disclosure.
[0050] FIG8 is a schematic structural diagram of an electronic device provided in Example 4 of the present disclosure. DETAILED DESCRIPTION
[0051] The present disclosure is further illustrated below by way of examples, but the present disclosure is not limited to the scope of the examples.
[0052] Example 1
[0053] 1 , the droplet screening method includes:
[0054] S1. Collect at least two electrical signals of the droplet to be tested according to a preset frequency, and each electrical signal of the droplet to be tested is obtained in a different analog-to-digital conversion signal channel.
[0055] Among them, a microfluidic sorting chip can be used when performing droplet screening. The droplets to be tested are arranged in a single row in the flow channel of the microfluidic sorting chip. When the spacing passes stably, the light source passes through the emission light path and is focused on the detection point of the microfluidic sorting chip. The light source and the emission light path are a light source module. Referring to Figure 2, the receiving light path installed at the other end uses a multi-channel photomultiplier tube (for example, a 7-channel photomultiplier tube) to convert the light signals of fluorescence, scattered light, and absorbed light into voltage signals. The multi-channel photomultiplier tubes are independently connected to an ADC module composed of multiple ADCs (analog-to-digital converters). Those skilled in the art will understand that the ADC module is composed of several analog-to-digital converters. The ADC module, the resolution of the ADC module, and the ADC acquisition frequency are determined by actual conditions. At least two electrical signals of the droplets to be tested are collected at a preset frequency, and each electrical signal of the droplets to be tested is obtained in different analog-to-digital conversion signal channels. For example, a droplet to be tested is collected at a frequency of 50KHZ, and the scattered light and fluorescence corresponding electrical signals of the droplet to be tested are collected, that is, the electrical signals corresponding to the scattered light and fluorescence are obtained in two analog-to-digital conversion signal channels. Among them, an ADC module composed of three ADCs is more common. If the ADC resolution is 12 and the reference voltage is 3.3V, the ADC reading is V adc , then the numerical conversion formula for the voltage level corresponding to the droplet is:
[0056] The unit is millivolt.
[0057] Background signal filtering can be completed through a control board connected to the ADC module, while simultaneously collecting characteristic values of multiple signals (different optical signals, or parameters of different bands of the same optical signal), including but not limited to peak value, average value, signal width, and performing signal calculations within and between channels to screen the droplets to be tested.
[0058] S2. Determine a target electrical signal in the electrical signal that meets an intensity threshold.
[0059] Among them, among the at least two electrical signals of the droplet to be tested collected at a preset frequency in different analog-to-digital conversion signal channels, the electrical signal that meets the intensity threshold for the first time in at least two analog-to-digital conversion signal channels is obtained as the starting point, until there is an electrical signal in one analog-to-digital conversion signal channel that does not meet the intensity threshold as the end point, and the electrical signal that meets the intensity threshold from the starting point to the end point is the target electrical signal. For example, the electrical signals corresponding to the scattered light and fluorescence of the droplet to be tested are collected, and the electrical signals that meet the intensity threshold in the two analog-to-digital conversion signal channels are obtained as the target electrical signals. The intensity threshold of each analog-to-digital conversion signal channel is determined according to the biological characteristics of the droplet to be tested, and the voltage level is usually 500mV (millivolts) to 1100mV (millivolts). The electrical signal that meets the intensity threshold for the first time in the two analog-to-digital conversion signal channels is obtained as the starting point, until there is an electrical signal in one analog-to-digital conversion signal channel that does not meet the intensity threshold as the end point, and the electrical signal that meets the intensity threshold from the starting point to the end point is the target electrical signal.
[0060] S3. When the signal width of the target electrical signal meets the signal width threshold, extract features from the target electrical signal, determine whether the intra-channel features of the target electrical signal meet the first preset condition, and determine whether the inter-channel features of the target electrical signal meet the second preset condition; if the judgment result is yes, execute step S31.
[0061] S31 , determining that the droplet to be measured is a target droplet.
[0062] When the signal widths of the target electrical signals obtained in different analog-to-digital conversion signal channels all meet the signal width threshold, it is determined whether the intra-channel characteristics of the target electrical signals meet a first preset condition and whether the inter-channel characteristics of the target electrical signals meet a second preset condition. If so, the droplet to be detected is determined to be the target droplet. If not, the process proceeds to S32 and discards the droplet.
[0063] The intra-channel feature is the feature of the droplet to be measured corresponding to each analog-to-digital conversion signal channel, and the inter-channel feature is the feature of the droplet to be measured corresponding to each analog-to-digital conversion signal channel.
[0064] In this embodiment, at least two electrical signals from a droplet to be tested are collected at a preset frequency, and a target electrical signal that meets a strength threshold is determined. When the signal width of the target electrical signal meets the signal width threshold, features are extracted from the target electrical signal data. The determination of whether the droplet to be tested is the target droplet is made by determining whether the intra-channel features of the target electrical signal meet a first preset condition and whether the inter-channel features of the target electrical signal meet a second preset condition. This provides a variety of screening options for droplets to be tested, improving the flexibility and accuracy of screening droplets to be tested.
[0065] Optionally, the signal width is obtained by calculating the number of target electrical signals that meet the intensity threshold and the acquisition frequency.
[0066] The signal width is calculated as:
[0067] S w =(N-1)*T sample ;
[0068] Among them, S w is the signal width, N is the number of target electrical signals that meet the intensity threshold in an analog-to-digital conversion signal channel for a droplet to be tested, and T sample It is the collection interval of a droplet to be tested.
[0069] In this embodiment, by calculating the signal width, a variety of screening schemes are provided for screening the droplets to be tested, thereby improving the flexibility and accuracy of screening the droplets to be tested.
[0070] Optionally, referring to FIG3 , the step of determining whether the intra-channel feature of the target electrical signal satisfies a first preset condition in step S3 includes:
[0071] S301 : Obtain the average intensity and peak intensity of the target electrical signal that meets the signal width threshold.
[0072] The average value and peak value of the voltage level intensity of the target electrical signal that meets the signal width threshold in different analog-to-digital conversion signal channels are obtained. For example, there are N target electrical signal voltage levels that meet the intensity threshold in one analog-to-digital conversion signal channel for a droplet to be tested, which are numbered as X. i , X i+1 …X N-1 , X N , the one with the largest voltage level is the peak intensity of the target electrical signal (V 峰值 ), the calculation formula of the average voltage level intensity of the target electrical signal is:
[0073]
[0074] S302. When the intra-channel feature of the target electrical signal that meets the signal width threshold meets the first ratio threshold, the first preset condition is met. The intra-channel feature is the ratio of the average intensity to the peak intensity of the target electrical signal that meets the signal width threshold.
[0075] Intra-channel features (R i_ma ) is calculated as:
[0076]
[0077] When the characteristics in the channel meet the first ratio threshold, the first preset condition is met, and the first ratio threshold is set according to the biological characteristics of the droplet to be detected.
[0078] In this embodiment, by calculating the characteristics within the channel and determining whether the characteristics within the channel meet the first preset condition, a variety of screening schemes are provided for screening the droplets to be tested, thereby improving the flexibility and accuracy of screening the droplets to be tested.
[0079] Optionally, the inter-channel feature includes a first inter-channel feature and a second inter-channel feature. Referring to FIG4 , the step of determining whether the inter-channel feature of the target electrical signal satisfies a second preset condition in step S3 includes:
[0080] S303: Acquire a target electrical signal that meets a first preset condition and obtain an intensity peak.
[0081] S304. When the first inter-channel characteristic of the target electrical signal that meets the first preset condition meets the second ratio threshold, the second preset condition is met. The inter-channel characteristic is the ratio of the peak intensity of the target electrical signal that meets the first preset condition between each analog-to-digital conversion signal channel corresponding to the droplet to be measured.
[0082] For example, the peak values of the target electrical signal voltage levels of a droplet to be tested that meet the first preset condition in two analog-to-digital conversion signal channels are obtained respectively, and the first inter-channel feature (R b_mm ), the calculation formula is:
[0083] V max1 is the peak voltage level intensity of the target electrical signal that meets the first preset condition in an analog-to-digital conversion signal channel, V max2 The voltage level intensity peak of the target electrical signal that meets the first preset condition in another analog-to-digital conversion signal channel is satisfied. When the first inter-channel characteristic meets the second ratio threshold, the second preset condition is satisfied.
[0084] and / or, see Figure 5,
[0085] S303', obtaining a target electrical signal that meets a first preset condition and obtaining an average strength value;
[0086] S304', when the second inter-channel characteristic of the target electrical signal that meets the first preset condition meets the third ratio threshold, the second preset condition is met, and the inter-channel characteristic is the ratio of the average intensity of the target electrical signal that meets the first preset condition between each analog-to-digital conversion signal channel corresponding to the droplet to be measured.
[0087] For example, the average value of the target electrical signal voltage level of a droplet to be tested that meets the first preset condition in two analog-to-digital conversion signal channels is obtained respectively, and the second inter-channel feature (R b_aa ), the calculation formula is:
[0088] V avg1is the average value of the target electrical signal voltage level that meets the first preset condition in an analog-to-digital conversion signal channel, V avg2 The intensity average value of the target electrical signal voltage level that meets the first preset condition in another analog-to-digital conversion signal channel is satisfied. When the second inter-channel feature meets the third ratio threshold, the second preset condition is satisfied.
[0089] In this embodiment, by calculating the first inter-channel feature and the second inter-channel feature and determining whether the inter-channel feature meets the second preset condition, a variety of screening schemes are provided for screening the droplets to be tested, thereby improving the flexibility and accuracy of screening the droplets to be tested.
[0090] Optionally, the step of collecting at least two electrical signals of the droplet to be tested at a preset frequency includes:
[0091] The optical signal of the droplet to be tested is obtained by a photomultiplier tube, and the optical signal is collected at a preset frequency to obtain at least two electrical signals of the droplet to be tested. The optical signal includes at least one of a scattered light signal, a fluorescent signal and an absorbed light signal.
[0092] At least one of the fluorescence signal, scattered light signal, and absorbed light signal is selected for screening based on the different biological characteristics of the droplets to be tested. For example, the fluorescence signal and scattered light of a droplet to be tested may correspond to two analog-to-digital conversion signal channels, respectively. Alternatively, two wavelength bands of the fluorescence signal of a droplet to be tested may correspond to two analog-to-digital conversion signal channels. Different optical signals and corresponding wavelength bands can be selected based on the biological characteristics of the droplet to be tested.
[0093] In this embodiment, fluorescent signals, scattered light signals and absorbed light signals are selected according to the biological characteristics of the droplets to be tested, and different wavelength bands are selected, thereby providing a variety of screening schemes for screening the droplets to be tested, thereby improving the flexibility and accuracy of screening the droplets to be tested.
[0094] Optionally, step S2 further includes:
[0095] When at least two electrical signals from a droplet to be tested, collected at a preset frequency, first meet an intensity threshold, the electrical signals are collected into a data packet until the electrical signals fall below the intensity threshold. The data packet collects all electrical signals from the at least two electrical signals from a droplet to be tested, collected at a preset frequency, from the first time they meet the intensity threshold to the last time they meet the intensity threshold.
[0096] Among them, among the at least two electrical signals of the droplet to be tested collected and obtained at a preset frequency in different analog-to-digital conversion signal channels, the electrical signal that meets the voltage level intensity threshold for the first time in at least two analog-to-digital conversion signal channels is obtained as the starting point, until there is an electrical signal in an analog-to-digital conversion signal channel that does not meet the voltage level intensity threshold as the end point, and the electrical signals that meet the voltage level intensity threshold from the starting point to the end point are collected into a data packet. When the voltage level intensity threshold is met, the data is updated to the data packet, and the voltage level, voltage level peak and number of the electrical signal are updated at the same time. Those skilled in the art should understand that in addition to the voltage level, other parameters can also be used for measurement, and different parameters can be selected according to actual conditions. In addition to the data packet, the data can also be stored in a data table or a data link. Analyze the data stored in the data packet, data table or data link, and analyze the electrical signal.
[0097] In this embodiment, by collecting data packets, data support is provided for the subsequent process of screening the droplets to be tested, a variety of screening schemes are provided for screening the droplets to be tested, and the flexibility and accuracy of screening the droplets to be tested are improved.
[0098] Optionally, when a droplet is determined to be positive, the excitation electrode generates a high voltage level, deflecting the droplet and completing the screening. The pulse width is selected based on the liquid flow rate and the distance between the detection point and the electrode, and ranges from 500 to 4000 microseconds.
[0099] Optionally, a communication protocol is customized according to the actual screening scheme, and operations such as querying the number of screening droplets and starting and stopping the screening program are performed through the host computer.
[0100] In a practical example, see Figure 6, two signals from channels 1 and 3 are used for droplet screening:
[0101] 1) The aqueous phase and droplet-forming oil are loaded into a syringe, which is connected to a tube. The tube is inserted into the corresponding inlet of the microfluidic chip. The syringe is opened to allow the droplets to enter the microfluidic sorting chip.
[0102] 2) Adjust the light intensity and position of the light source so that the light spot is focused on the detection point; monitor the voltage level of the drip signal through an oscilloscope and control the signal peak voltage between 2V (volt-ampere) and 3V (volt-ampere) to ensure that the light intensity and position are suitable for the subsequent screening process.
[0103] 3) Adjust the syringe flow rate and capture the interval between two droplet signals using an oscilloscope so that the interval between the two droplet signals is approximately 4 milliseconds, that is, the flow rate is controlled at 250 droplets / second.
[0104] 4) The acquisition frequency has been set to 100K Hz in the software, that is, the acquisition interval T sample =10 microseconds.
[0105] 5) Set parameters.
[0106] Channel 1 parameters: voltage level intensity threshold V thr_1 =850mV, average voltage level strength V avg_1 =1250mV, voltage level peak strength V max_1 =1600mV, signal width threshold S w_1 =120;
[0107] Channel 3 parameters: voltage level intensity threshold V thr_3 =750mV, average voltage level strength V avg_3 =1100mV, voltage level peak value V max_3 =1300mV, signal width threshold S w_3 =120.
[0108] First channel characteristics:
[0109] 6) Start the timer and begin screening.
[0110] 7) In-channel judgment: when the voltage level of the electrical signal of channel 1 is ≥V thr_1 and the electrical signal voltage level of channel 3 ≥ V thr_3 If it is the first time that the signal is satisfied, it is recorded as the signal starting point and the data is stored. thr_1 And the electrical signal voltage level of channel 3 is less than V thr_3 When the signal width of channel 1 is ≥ S w_1 And channel 3 signal width ≥ S w_3 If both conditions are met, proceed to step 8) to determine the characteristic conditions within the channel; otherwise, clear the cache and proceed to the next droplet screening.
[0111] 8) Inter-channel judgment:
[0112] Channel 1 judgment: the average value of the electrical signal voltage level ≥ V avg_1 , peak value of electrical signal voltage level ≥V max_1 ;
[0113] Channel 3 judgment: average value of electrical signal voltage level ≥ V avg_3 , peak value of electrical signal voltage level ≥V max_3 If the two channel characteristic conditions are met, proceed to step 9) to determine the inter-channel characteristic conditions; otherwise, clear the cache and proceed to the next droplet screening.
[0114] 9) Calculate the voltage level intensity peak ratio ≥ R b_mmIf the conditions are met, the electrode is excited to deflect the micro-droplet, thus realizing the screening of a droplet; otherwise, the cache is cleared and the next droplet screening is carried out.
[0115] 10) Repeat steps 7), 8), and 9 to achieve automatic screening of microdroplets. During the screening process, the host computer can query the number of screened droplets every 1 second according to the communication protocol to facilitate statistics and analysis of the droplet distribution.
[0116] Example 2
[0117] Referring to Figure 7, the droplet screening system includes:
[0118] The acquisition module 101 is used to collect at least two electrical signals of the droplet to be tested according to a preset frequency, and each electrical signal of the droplet to be tested is acquired in a different analog-to-digital conversion signal channel.
[0119] Among them, when performing droplet screening, a microfluidic sorting chip can be used. The droplets to be tested are arranged in a single row in the flow channel of the microfluidic sorting chip. When the spacing is stable, the light source passes through the emission light path and is focused on the detection point of the microfluidic sorting chip. Referring to Figure 2, the receiving light path installed at the other end uses a multi-channel photomultiplier tube (for example, a 7-channel photomultiplier tube) to convert the light signals of fluorescence, scattered light, and absorbed light into voltage signals. The multi-channel photomultiplier tube is independently connected to an ADC module composed of multiple ADCs (analog-to-digital converters). Those skilled in the art will understand that the ADC module is composed of several analog-to-digital converters. The ADC module, the resolution of the ADC module, and the ADC acquisition frequency are determined by actual conditions. At least two electrical signals of the droplets to be tested are collected at a preset frequency, and each electrical signal of the droplets to be tested is obtained in different analog-to-digital conversion signal channels. For example, a droplet to be tested is collected at a frequency of 50KHZ, and the electrical signals corresponding to the scattered light and fluorescence of the droplet to be tested are collected, that is, the electrical signals corresponding to the scattered light and fluorescence are obtained in two analog-to-digital conversion signal channels. Among them, the ADC module composed of three ADCs is more common. If the ADC resolution is 12, the reference voltage is 3.3V, and the ADC reading is V adc , then the numerical conversion formula for the voltage level corresponding to the droplet is:
[0120] The unit is millivolt.
[0121] The determination module 102 is configured to determine a target electrical signal in the electrical signal that meets a strength threshold.
[0122] Among them, among the at least two electrical signals of the droplet to be tested collected at a preset frequency in different analog-to-digital conversion signal channels, the electrical signal that meets the intensity threshold for the first time in at least two analog-to-digital conversion signal channels is obtained as the starting point, until there is an electrical signal in one analog-to-digital conversion signal channel that does not meet the intensity threshold as the end point, and the electrical signal that meets the intensity threshold from the starting point to the end point is the target electrical signal. For example, the electrical signals corresponding to the scattered light and fluorescence of the droplet to be tested are collected, and the electrical signals that meet the intensity threshold in the two analog-to-digital conversion signal channels are obtained as the target electrical signals. The intensity threshold of each analog-to-digital conversion signal channel is determined according to the biological characteristics of the droplet to be tested, and the voltage level is usually 500mV (millivolts) to 1100mV (millivolts). The electrical signal that meets the intensity threshold for the first time in the two analog-to-digital conversion signal channels is obtained as the starting point, until there is an electrical signal in one analog-to-digital conversion signal channel that does not meet the intensity threshold as the end point, and the electrical signal that meets the intensity threshold from the starting point to the end point is the target electrical signal.
[0123] The judgment module 103 is used to extract features from the target electrical signal data when the signal width of the target electrical signal meets the signal width threshold, and to judge whether the intra-channel features of the target electrical signal meet the first preset condition and whether the inter-channel features of the target electrical signal meet the second preset condition.
[0124] If the conditions are met, the droplet to be tested is determined to be the target droplet.
[0125] The intra-channel feature is the feature of the droplet to be measured corresponding to each analog-to-digital conversion signal channel, and the inter-channel feature is the feature of the droplet to be measured corresponding to each analog-to-digital conversion signal channel.
[0126] When the signal widths of the target electrical signals obtained in different analog-to-digital conversion signal channels all meet the signal width threshold, it is determined whether the intra-channel characteristics of the target electrical signal meet the first preset condition and whether the inter-channel characteristics of the target electrical signal meet the second preset condition. If so, the droplet to be tested is determined to be the target droplet.
[0127] The intra-channel feature is the feature of the droplet to be measured corresponding to each analog-to-digital conversion signal channel, and the inter-channel feature is the feature of the droplet to be measured corresponding to each analog-to-digital conversion signal channel.
[0128] In this embodiment, at least two electrical signals from a droplet to be tested are collected at a preset frequency, and a target electrical signal that meets a strength threshold is determined. When the signal width of the target electrical signal meets the signal width threshold, features are extracted from the target electrical signal data. The determination of whether the droplet to be tested is the target droplet is made by determining whether the intra-channel features of the target electrical signal meet a first preset condition and whether the inter-channel features of the target electrical signal meet a second preset condition. This provides a variety of screening options for droplets to be tested, improving the flexibility and accuracy of screening droplets to be tested.
[0129] Optionally, the signal width is obtained by calculating the number of target electrical signals that meet an intensity threshold and an acquisition frequency.
[0130] Optionally, when determining whether the intra-channel feature of the target electrical signal meets a first preset condition, the determination module is specifically configured to:
[0131] Obtaining an average intensity value and a peak intensity value of the target electrical signal that meets a signal width threshold;
[0132] When the in-channel feature of the target electrical signal that meets the signal width threshold meets the first ratio threshold, the first preset condition is met;
[0133] The intra-channel feature is the ratio of the average intensity to the peak intensity of the target electrical signal that meets the signal width threshold.
[0134] Optionally, the inter-channel feature includes a first inter-channel feature and a second inter-channel feature. When determining whether the inter-channel feature of the target electrical signal meets a second preset condition, the determination module is specifically configured to:
[0135] Acquire the target electrical signal that meets the first preset condition and obtain the intensity peak;
[0136] When the first inter-channel feature of the target electrical signal that meets the first preset condition meets the second ratio threshold, the second preset condition is met;
[0137] The inter-channel characteristic is a ratio of the peak values of the target electrical signals corresponding to the analog-to-digital conversion signal channels that meet a first preset condition.
[0138] and / or,
[0139] Acquire the target electrical signal that meets the first preset condition and obtain an average strength value;
[0140] When the second inter-channel feature of the target electrical signal that meets the first preset condition meets the third ratio threshold, the second preset condition is met;
[0141] The inter-channel characteristic is a ratio of the average intensity of the target electrical signal corresponding to each of the analog-to-digital conversion signal channels that meets a first preset condition.
[0142] Optionally, the acquisition module is specifically configured to:
[0143] Acquire an optical signal of the droplet to be measured by a photomultiplier tube, collect the optical signal at a preset frequency and acquire at least two electrical signals of the droplet to be measured;
[0144] The optical signal includes at least one of a scattered light signal, a fluorescent signal and an absorbed light signal.
[0145] Optionally, the determining module is specifically configured to:
[0146] When at least two electrical signals of the droplet to be tested collected at a preset frequency meet an intensity threshold for the first time, the electrical signals are collected into a data packet;
[0147] until the electrical signal is lower than a strength threshold;
[0148] The data packet collects all electrical signals of at least two electrical signals of the droplet to be tested from the first time the signal meets the intensity threshold to the last time the signal meets the intensity threshold according to a preset frequency.
[0149] As for the system embodiment, since it basically corresponds to the method embodiment, the relevant parts can be referred to the partial description of the method embodiment. The system embodiment described above is only illustrative, in which the units described as separate components may or may not be physically separated, and the components shown as units may or may not be physical units, that is, they may be located in one place or distributed across multiple network units. Some or all of the modules can be selected according to actual needs to achieve the purpose of the solution of the present invention. Those of ordinary skill in the art can understand and implement it without expending creative work.
[0150] Example 3
[0151] The present disclosure provides a microfluidic sorting chip, which includes a sorting component and at least two branch channels. When a droplet to be tested is determined to be a target droplet by the droplet screening method of the above-mentioned embodiment 1, the sorting component is used to receive a deflection instruction and sort the droplet to be tested into a corresponding branch channel.
[0152] As shown in Figure 2, the aqueous and oil phases enter the microfluidic chip through inlets 1, 2, and 3, forming droplets. When a droplet is identified as a target droplet, the sorting component receives a deflection command and deflects the droplet via an excitation electrode. Finally, the target droplet and waste droplets flow out of the branch channel through outlets 1 and 2.
[0153] In this embodiment, the use of a microfluidic sorting chip makes the amount of reagent used in the microdroplets very small, thereby greatly reducing the cost.
[0154] Example 4
[0155] This embodiment provides an electronic device, which can be expressed in the form of a computing device (for example, a server device), including a memory, a processor, and a computer program stored in the memory and executable on the processor. When the processor executes the computer program, the droplet screening method provided in Example 1 can be implemented.
[0156] FIG8 shows a schematic diagram of the hardware structure of this embodiment. As shown in FIG8 , the electronic device 9 specifically includes:
[0157] At least one processor 91, at least one memory 92, and a bus 93 for connecting different system components (including the processor 91 and the memory 92), wherein:
[0158] The bus 93 includes a data bus, an address bus, and a control bus.
[0159] The memory 92 includes a volatile memory, such as a random access memory (RAM) 921 and / or a cache memory 922 , and may further include a read-only memory (ROM) 923 .
[0160] Memory 92 also includes a program / utility 925 having a set (at least one) of program modules 924, such program modules 924 including but not limited to: an operating system, one or more application programs, other program modules, and program data, each of which or some combination may include an implementation of a network environment.
[0161] The processor 91 executes various functional applications and data processing by running computer programs stored in the memory 92, such as the droplet screening method provided in Example 1 of the present disclosure.
[0162] The electronic device 9 can further communicate with one or more external devices 94. Such communication can be performed via an input / output (I / O) interface 95. Furthermore, the electronic device 9 can communicate with one or more networks (e.g., a local area network (LAN), a wide area network (WAN), and / or a public network, such as the Internet) via a network adapter 96. The network adapter 96 communicates with other modules of the electronic device 9 via a bus 93. It should be understood that, although not shown in the figures, other hardware and / or software modules can be used in conjunction with the electronic device 9, including but not limited to microcode, device drivers, redundant processors, external disk drive arrays, RAID (RAID) systems, tape drives, and data backup storage systems.
[0163] It should be noted that although several units / modules or sub-units / modules of the electronic device are mentioned in the above detailed description, this division is merely exemplary and not mandatory. In fact, depending on the embodiment of the present application, the features and functions of two or more units / modules described above can be embodied in one unit / module. Conversely, the features and functions of one unit / module described above can be further divided and embodied by multiple units / modules.
[0164] Example 5
[0165] This embodiment provides a computer-readable storage medium having a computer program stored thereon. When the program is executed by a processor, the droplet screening method provided in Example 1 is implemented.
[0166] Specifically, the readable storage medium may include, but is not limited to, portable disks, hard disks, random access memories, read-only memories, erasable programmable read-only memories, optical storage devices, magnetic storage devices, or any suitable combination thereof.
[0167] In a possible implementation, the present disclosure may also be implemented in the form of a program product, which includes program code. When the program product is run on a terminal device, the program code is used to enable the terminal device to execute the droplet screening method provided in Example 1.
[0168] The program code for executing the present disclosure may be written in any combination of one or more programming languages, and may be executed entirely on the user device, partially on the user device, as a standalone software package, partially on the user device and partially on a remote device, or entirely on the remote device.
[0169] While specific embodiments of the present disclosure have been described above, those skilled in the art will appreciate that these are merely illustrative and that the scope of protection of the present disclosure is defined by the appended claims. Those skilled in the art may make various changes or modifications to these embodiments without departing from the principles and essence of the present disclosure, and such changes and modifications are intended to fall within the scope of protection of the present disclosure.
Claims
1. A droplet screening method, characterized in that: The method comprises: Collecting at least two electrical signals of the droplet to be tested at a preset frequency, wherein each electrical signal of the droplet to be tested is obtained in a different analog-to-digital conversion signal channel; Determine a target electrical signal in the electrical signal that meets a strength threshold; When the signal width of the target electrical signal meets the signal width threshold, extracting features from the target electrical signal data, determining whether the intra-channel features of the target electrical signal meet a first preset condition, and determining whether the inter-channel features of the target electrical signal meet a second preset condition; If yes, the droplet to be tested is determined to be a target droplet; The intra-channel feature is a feature of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels, and the inter-channel feature is a feature of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels.
2. The droplet screening method according to claim 1, characterized in that: The signal width is obtained by calculating the number of target electrical signals that meet the intensity threshold and the acquisition frequency.
3. The droplet screening method according to claim 1, characterized in that: The step of determining whether the intra-channel feature of the target electrical signal meets a first preset condition comprises: Obtaining an average intensity value and a peak intensity value of the target electrical signal that meets a signal width threshold; When the in-channel feature of the target electrical signal that meets the signal width threshold meets the first ratio threshold, the first preset condition is met; The intra-channel feature is the ratio of the average intensity of the target electrical signal that meets the signal width threshold to the peak intensity.
4. The droplet screening method according to claim 3, characterized in that: The inter-channel feature includes a first inter-channel feature and a second inter-channel feature, and the step of determining whether the inter-channel feature of the target electrical signal meets a second preset condition includes: Acquire the target electrical signal that meets the first preset condition and obtain the intensity peak value; When the first inter-channel feature of the target electrical signal that satisfies the first preset condition satisfies the second ratio threshold, the second preset condition is satisfied; The inter-channel characteristic is the ratio of the peak intensity of the target electrical signal between the droplets to be tested corresponding to each of the analog-to-digital conversion signal channels that meets the first preset condition; and / or, Acquire the target electrical signal that meets the first preset condition and obtain an average strength value; When the second inter-channel feature of the target electrical signal that satisfies the first preset condition satisfies the third ratio threshold, the second preset condition is satisfied; The inter-channel characteristic is the ratio of the average intensity of the target electrical signal between the droplets to be tested corresponding to each of the analog-to-digital conversion signal channels that meets the first preset condition.
5. The droplet screening method according to claim 1, characterized in that: The step of collecting at least two electrical signals of the droplet to be tested according to a preset frequency comprises: Acquire the optical signal of the droplet to be detected by a photomultiplier tube, collect the optical signal by a preset frequency and acquire at least two electrical signals of the droplet to be detected; The optical signal includes at least one of a scattered light signal, a fluorescent signal and an absorbed light signal.
6. The droplet screening method according to claim 1, characterized in that: The step of determining a target electrical signal in the electrical signal that meets a strength threshold further includes: When at least two electrical signals of the droplet to be tested collected at a preset frequency meet an intensity threshold for the first time, the electrical signals are collected into a data packet; until the electrical signal is below a strength threshold; The data packet collects all electrical signals of at least two electrical signals of the droplet to be tested from the first time the signal meets the intensity threshold to the last time the signal meets the intensity threshold according to a preset frequency.
7. A microfluidic sorting chip, characterized in that: The microfluidic sorting chip includes a sorting component and at least two branch channels. When the droplet to be tested is determined to be a target droplet by the droplet screening method described in any one of claims 1 to 6, the sorting component is used to receive a deflection instruction to sort the droplet to be tested into a corresponding branch channel.
8. A droplet screening system, characterized in that: The system comprises: An acquisition module, used for acquiring at least two electrical signals of the droplet to be tested according to a preset frequency, wherein each electrical signal of the droplet to be tested is acquired in a different analog-to-digital conversion signal channel; A determination module, used to determine a target electrical signal in the electrical signal that meets a strength threshold; A judgment module, configured to extract features from the target electrical signal data when the signal width of the target electrical signal meets a signal width threshold, judge whether the intra-channel features of the target electrical signal meet a first preset condition, and judge whether the inter-channel features of the target electrical signal meet a second preset condition; If yes, the droplet to be tested is determined to be a target droplet; The intra-channel feature is a feature of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels, and the inter-channel feature is a feature of the droplet to be measured corresponding to each of the analog-to-digital conversion signal channels.
9. An electronic device comprising a memory, a processor, and a computer program stored in the memory and used to run on the processor, characterized in that: When the processor executes the computer program, the droplet screening method according to any one of claims 1 to 6 is implemented.
10. A computer-readable storage medium having a computer program stored thereon, characterized in that: When the computer program is executed by a processor, the droplet screening method according to any one of claims 1 to 6 is implemented.