Method for quantitatively distinguishing signals of magnetic nanoparticles inside and outside cells based on magnetic particle spectrum

Abstract

The invention belongs to the field of magnetic nanoparticle imaging, particularly relates to a method for quantitatively distinguishing signals of magnetic nanoparticles inside and outside cells based on a magnetic particle spectrum, electronic equipment and a computer readable storage medium, and aims to solve the problem that the influence of free SPIO after death of labeled cells cannot be eliminated on a signal level without considering the influence of cell survival conditions. The method comprises the following steps: according to the periodicity of a magnetic nanoparticle signal, calculating an attenuation ratio of odd-order harmonic amplitudes in MPS, indicating the aggregation or dispersion state of SPIO in and out of cells, constructing a distribution model, calculating a proportionality coefficient, and correcting and obtaining an intracellular SPIO signal of an unwashed labeled cell sample, and extraction and separation of intracellular and extracellular SPIO signals are realized. According to the method, SPIO signals in the cells are separated and extracted, the survival labeled cells are traced through MPI, and MPI stem cell tracing is more accurate and reliable in clinical application.

Description

Background Art

[0002] Magnetic Particle Imaging (MPI) is an imaging technology based on superparamagnetic iron oxide nanoparticles (SPIOs). It generates images by measuring the response of magnetic nanoparticles in a specific magnetic field. The imaging principle of MPI is based on the nonlinear magnetization curve of the Langevin paramagnetic law. By constructing a gradient field to generate a field-free point (FFP) or a field-free line (FFL), the magnetic nanoparticles (SPIO) in FFP or FFL have not reached the magnetic saturation state and can produce a nonlinear magnetization response to the excitation magnetic field. The signal voltage is detected in the receiving coil, and the voltage signal is distributed to each position for reconstruction, so that the spatial distribution information of the magnetic particles, namely the MPI signal, can be obtained. Since the body tissue has diamagnetic properties and does not generate any interference signals, MPI does not display anatomical structures.

[0003] Currently, the research on MPI is in its infancy. It has the advantages of high sensitivity, no radiation, and no need for external radiation sources, and has broad application potential in the field of biomedical imaging. Compared with traditional magnetic resonance imaging (MRI), MPI can provide higher temporal resolution and contrast, and is particularly suitable for monitoring dynamic processes such as blood flow, cell migration, and drug distribution. In addition, MPI also shows great potential in quantitative analysis, and can accurately measure the concentration of magnetic particles in tissues, providing important information for disease diagnosis and treatment monitoring.

[0004] However, in the current MPI-based quantitative stem cell tracing studies, most studies have not considered the impact of cell survival: whether the label will persist after cell death, and whether the cell membrane will rupture after cell death, allowing the SPIO label to be transferred to other cells, causing signal interference. Summary of the invention

[0005] In order to solve the above-mentioned problem in the prior art, that is, the prior art does not consider the influence of cell survival, and cannot exclude the influence of free SPIO after the death of labeled cells at the signal level, resulting in signal interference, and further resulting in the inability to accurately obtain the SPIO signal in the living cells of the unwashed labeled cell sample, the first aspect of the present invention proposes a method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy, which is used to extract and separate intracellular and extracellular SPIO signals, and then obtain the SPIO signal in the cells of the unwashed labeled cell sample. The method comprises the following steps: Obtaining a response voltage signal U1 of the washed labeled cell sample under stimulation; Obtaining a response voltage signal U2 of the sample with ruptured cell membrane under excitation; Obtaining a response voltage signal U3 of an unwashed labeled cell sample under stimulation; Perform Fourier transformation on U1 to obtain the magnetic particle spectrum of SPIO in the cell, and then calculate the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K1; Performing Fourier transformation on U2 to obtain the magnetic particle spectrum of extracellular SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K2; Based on the K1 and the K2, constructing a distribution model of SPIO inside and outside the cell; Performing Fourier transformation on the U3 to obtain the magnetic particle spectrum of the unwashed labeled cell sample SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K3; In combination with the distribution model and K3, a proportional coefficient between intracellular SPIO and extracellular SPIO in the unwashed labeled cell sample is solved; The U3 is corrected according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample.

[0006] In some preferred embodiments, the ratio of the difference between the amplitudes of adjacent odd-order harmonics is calculated as: (A3-A5) / (A5-A7), wherein A3 is the amplitude of the third harmonic, A5 is the amplitude of the fifth harmonic, and A7 is the amplitude of the seventh harmonic.

[0007] In some preferred embodiments, the distribution model of SPIO inside and outside the cell is: ; In the formula, For the labeled cell samples after washing (A5-A7), is the sample with ruptured cell membrane (A5-A7), m is the mixed sample, α is the proportion of the labeled cell sample after washing in the mixed sample, is the proportion of samples with ruptured cell membranes in the mixed samples, It is the ratio of the odd harmonic differences of the mixed samples (A3-A5) / (A5-A7).

[0008] In some preferred embodiments, the proportionality coefficient is calculated as follows: Transform the distribution model equation and solve for the proportionality coefficient α: ; For the unwashed labeled cell sample to be tested, the ratio of its odd harmonic differences K 3 Bring in , the ratio α of the intracellular SPIO signal in the unwashed labeled cell sample to the total signal U3 is obtained as: .

[0009] In some preferred embodiments, the U3 is corrected according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample, and the method is: Step S71, obtaining a standard unit signal u1 generated by SPIO in the cell, wherein u1 is U1 divided by the total number of cells in the labeled cell sample after washing; Step S72, obtaining a standard unit signal u2 generated by extracellular SPIO, wherein u2 is U2 divided by the total number of cells in the sample with ruptured cell membranes; Step S73, combining the u1 and the u2, calibrating the U3 signal of the unwashed labeled cell sample: ; In the formula, is the signal generated by intracellular SPIO in the unwashed labeled cell sample, and n is the number of cells in the unwashed labeled cell sample.

[0010] In some preferred embodiments, after obtaining the SPIO signal in the cells of the unwashed labeled cell sample, the method further includes quickly correcting the obtained SPIO signal by: Fitting the U1 harmonic curve and K1 curve of SPIO particle incubated cells R at different magnetic nanoparticle incubation concentrations; According to the voltage response signal U2 of SPIO outside the cell and the voltage response signal U 2-pbs The relationship between features, constructing a relational expression; Fitting the U of SPIO diluted samples with different concentrations 2-pbs Harmonic curve, K 2-pbs Curve, and obtain U2 harmonic curve and K2 curve based on the relationship; According to the SPIO concentration of the incubated cells, K1 and K2 are respectively found on the K1 curve and the K2 curve, and Δ is calculated according to the corresponding harmonic amplitudes on the U1 harmonic curve and the U2 harmonic curve. 1 , Δ 2 ; The K2, K1, Δ 1 , Δ 2, the measured cell sample signals U3 and K3 of the unknown SPIO distribution state are substituted into the proportional coefficient α to solve the formula, and then the proportional coefficient of SPIO inside and outside the cell is obtained; Calculating the signal measurement deviation based on the relationship and the proportionality coefficient of SPIO inside and outside the cell; Based on the measured deviation of the signal, the measured cell sample signal U3 of the unknown SPIO distribution state is corrected to obtain the SPIO signal in the cell .

[0011] In some preferred embodiments, the relationship is: ; The measured deviation of the signal is: ; The intracellular SPIO signal U in : .

[0012] In a second aspect of the present invention, a system for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy is used to extract and separate intracellular and extracellular SPIO signals, thereby obtaining intracellular SPIO signals of unwashed labeled cell samples, and the system comprises: The MPI signal detection instrument is used to excite and scan the washed labeled cell samples, the samples with ruptured cell membranes, and the unwashed labeled cell samples to obtain response voltage signals U1, U2, and U3; The central processing unit includes a CPU and a GPU; the central processing unit is used to perform Fourier transform on U1 to obtain the magnetic particle spectrum of SPIO in the cell, and then calculate the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K1; Performing Fourier transformation on U2 to obtain the magnetic particle spectrum of extracellular SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K2; Based on the K1 and the K2, constructing a distribution model of SPIO inside and outside the cell; Performing Fourier transformation on the U3 to obtain the magnetic particle spectrum of the unwashed labeled cell sample SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K3; In combination with the distribution model and K3, a proportional coefficient between intracellular SPIO and extracellular SPIO in the unwashed labeled cell sample is solved; The U3 is corrected according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample.

[0013] According to a third aspect of the present invention, an electronic device is provided, comprising: At least one processor; and a memory communicatively connected to at least one of the processors; wherein the memory stores instructions executable by the processor, and the instructions are used to be executed by the processor to implement the method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy.

[0014] In a fourth aspect of the present invention, a computer-readable storage medium is proposed, wherein the computer-readable storage medium stores computer instructions, and the computer instructions are used to be executed by the computer to implement the above-mentioned method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy.

[0015] Beneficial effects of the present invention: 1. The constructed SPIO distribution model inside and outside cells is used to extract the ratio of the magnetic particle spectrum (A3-A5) / (A5-A7) of standard intracellular and extracellular SPIO. Without the aid of other tests, the intensity ratio of the response voltage signal generated by intracellular SPIO and extracellular SPIO is obtained. The response voltage signal generated by intracellular SPIO is separated and extracted at the signal level, and the proportional coefficient is calculated and corrected to obtain the SPIO signal inside the cell of the unwashed labeled cell sample. The SPIO signal inside the cell can be separated and extracted to achieve accurate MPI tracing of surviving labeled cells, which makes up for the problem that the influence of free SPIO cannot be excluded at the signal level in the current MPI cell tracing research. 2. It can also promote the improvement of MPI detection sensitivity. The method proposed in this study can realize the detection of even smaller amounts of SPIO signals in cells, which is in line with the development needs and direction of precision medicine. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Other features, objects and advantages of the present application will become more apparent by reading the detailed description of non-limiting embodiments made with reference to the following drawings: Figure 1 It is the SPIO signal spectrum diagram inside the living cells and outside the dead cells of the present invention; Figure 2 This is a difference diagram of the living cells outside the dead cells under different incubation concentrations of the present invention; Figure 3 This is a spectrum diagram of the extracellular SPIO magnetic particles of the present invention; Figure 4 This is a spectrum diagram of SPIO magnetic particles in cells of the present invention; Figure 5 is a standard curve diagram of the amplitude of odd-order harmonics at different incubation concentrations of the present invention; Figure 6is a standard curve diagram of K1 and K2 at different incubation concentrations of the present invention; Figure 7 It is a signal diagram of SPIO magnetic particles in cells extracted from the mixed sample signal of the present invention; Figure 8 It is a flow chart of the steps of the method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy of the present invention. DETAILED DESCRIPTION

[0017] The present application will be further described in detail below in conjunction with the accompanying drawings and embodiments. It is to be understood that the specific embodiments described herein are only used to explain the relevant invention, rather than to limit the invention. It is also necessary to explain that, for ease of description, only the parts related to the relevant invention are shown in the accompanying drawings.

[0018] It should be noted that, in the absence of conflict, the embodiments and features in the embodiments of the present application can be combined with each other. The present application will be described in detail below with reference to the accompanying drawings and in combination with the embodiments.

[0019] In order to more clearly illustrate the method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy of the present invention, the following is combined with Figures 1 to 8 Each step in the embodiment of the present invention is described in detail.

[0020] In the experiment, it was found that under the same measurement environment, the signal of SPIO tag in intact living cells is different from that after cell membrane rupture and death. Specifically, the amplitude of odd harmonics in MPS is different. The amplitude attenuation of odd harmonics of SPIO signal after cell membrane rupture and death is greater than that of SPIO signal in living cells. Figure 1 , showing the signal spectrum characteristics of MSC cells incubated with Synomag 50nm magnetic nanoparticle solution during survival and cell membrane rupture and death, see Figure 2, showing the difference in (A3-A5) / (A5-A7) between the viable cells and the dead cells under different incubation concentrations. Table 1 shows the difference between the SPIO signals (A3-A5) / (A5-A7) of the dead cells and the SPIO signals (A3-A5) / (A5-A7) of the viable cells in the 12 measurement results. Based on this, the first embodiment of the present invention proposes a method for quantitatively distinguishing the signals of magnetic nanoparticles inside and outside the cell based on magnetic particle spectroscopy. The nonlinear magnetization response of SPIO in an alternating magnetic field is used to transform the response voltage signal into the frequency domain, and the odd harmonic attenuation (A3-A5) / (A5-A7) in its MPS is analyzed. This ratio can indicate the aggregation or dispersion state of SPIO inside and outside the cell. The (A3-A5) / (A5-A7) characteristics of the SPIO signals inside and outside the cell are used to construct a distribution model of SPIO inside and outside the cell, so as to achieve the distinction of SPIO signals inside and outside the cell and the extraction of SPIO signals inside the cell.

[0021] Table 1 concentration 10 μg / mL 25 μg / mL 50 μg / mL 75 μg / mL 100 μg / mL 200 μg / mL -2.4369497 6.45305542 1.84719034 3.18464156 0.23368361 0.5155164 -2.0462797 0.76660302 1.86137282 0.04648958 0.80739233 0.77167248 See also Figure 8 , as follows: Step S1, obtaining a response voltage signal U1 of the washed labeled cell sample under stimulation; In this embodiment, the process of obtaining the response voltage signal U1 under excitation is as follows: the washed labeled cell sample (i.e., the intracellular SPIO standard) is placed in the MPI signal detection instrument and placed at the center of the FOV for scanning measurement to obtain the response voltage signal U1 under excitation, wherein the preparation process of the washed labeled cell sample is as follows: first, a SPIO culture medium dilution solution is prepared, and the standard requires an iron concentration of 50 μg / mL. It is noted that in actual configuration, the total volume is not limited, and the area of ​​the culture dish or culture bottle is used as the standard, and the iron concentration is also not limited; secondly, the cells are incubated with the SPIO culture medium dilution solution standard, and the incubation time is calculated according to one cell cycle of the labeled cells; finally, after the incubation is completed, the labeled cells are centrifuged, the SPIO-containing culture medium is removed, and the SPIO particles outside the cells are washed twice with PBS, the cells marked with SPIO are digested, and 1 ml of culture medium without SPIO is added to resuspend the cells as the intracellular SPIO standard; Step S2, obtaining a response voltage signal U2 of the sample with ruptured cell membrane under stimulation; In this embodiment, the process of obtaining the response voltage signal U2 of the sample with ruptured cell membrane under excitation is as follows: the sample with ruptured cell membrane (i.e., the extracellular SPIO standard) is placed in the MPI signal detection instrument and placed at the center of the FOV for scanning measurement, and the scanning measurement is performed to obtain the response voltage signal U2 under excitation, wherein the sample preparation process of ruptured cell membrane is as follows: the cell membrane of the intracellular SPIO standard prepared in step S1 is ruptured to release SPIO particles into the solution as the extracellular SPIO standard; Step S3, obtaining a response voltage signal U3 of the unwashed labeled cell sample under stimulation; In this embodiment, the process of obtaining the response voltage signal U3 of the unwashed labeled cell sample under excitation is as follows: the unwashed labeled cell sample is placed in the MPI signal detection instrument and placed in the center of the FOV, and a scanning measurement is performed to obtain the response voltage signal U3 under excitation, wherein the preparation process of the unwashed labeled cell sample is: the SPIO standard in the cell is re-inoculated into the culture dish, and after incubation for 48 hours, the old culture medium is sucked out for standby use, and after the cell digestion and centrifugation, the cells are resuspended with the old culture medium and directly used as the unwashed labeled cell sample; Step S4, performing Fourier transform on U1 to obtain the magnetic particle spectrum of SPIO in the cell, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, and recording it as K1; Performing Fourier transformation on U2 to obtain the magnetic particle spectrum of extracellular SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K2; In this embodiment, the calculation method of K1 and K2 is: Step S41: Obtain the U1, convert the U1 from the time domain to the frequency domain using Fourier transform, and obtain the magnetic particle spectrum of SPIO in the cell, see Figure 3 ; Obtain the U2, convert the U2 from the time domain to the frequency domain using Fourier transform, and obtain the magnetic particle spectrum of the extracellular SPIO, see Figure 4 ; Step S42: Since the amplitude of the high-order harmonics is too small, only the attenuation speed differences of the amplitudes of the 3rd, 5th and 7th harmonics (i.e. A3, A5, A7) are compared, and the ratio of the odd-order harmonic differences of the intracellular SPIO magnetic particle spectrum (A3-A5) / (A5-A7) is calculated and recorded as K1; Since the amplitude of high-order harmonics is too small, only the attenuation speed difference of the amplitude of the 3rd, 5th and 7th harmonics (i.e. A3, A5, A7) is compared, and the ratio of the odd-order harmonic difference of the extracellular SPIO magnetic particle spectrum is calculated (A3-A5) / (A5-A7), which is recorded as K2; Step S5, constructing a distribution model of SPIO inside and outside the cell based on K1 and K2; In this example, the model is used to describe the changes in the sample (A3-A5) / (A5-A7) ratio caused by the processes of SPIO being engulfed into cells and SPIO being released outside the cells again due to cell death and other reasons. The distribution model is: ; In the formula, For the labeled cell samples after washing (A5-A7), is the sample with ruptured cell membrane (A5-A7), m is the mixed sample, α is the proportion of the labeled cell sample after washing in the mixed sample, is the proportion of samples with ruptured cell membranes in the mixed samples, is the ratio of the odd harmonic differences of the mixed samples (A3-A5) / (A5-A7); Step S6: Perform Fourier transform on U3 to obtain the magnetic particle spectrum of the unwashed labeled cell sample SPIO, and then calculate the ratio of the difference between the amplitudes of adjacent odd harmonics, and record it as K3 In this embodiment, the process of calculating K3 is as follows: obtain U3, convert U3 from the time domain to the frequency domain using Fourier transform, obtain the magnetic particle spectrum of the unwashed labeled cell sample, see Figure 4 ; Since the amplitude of the high-order harmonics is too small, only the attenuation speed difference of the amplitude of the 3rd, 5th and 7th harmonics (i.e. A3, A5, A7) is compared, and the ratio of the odd-order harmonic difference of the SPIO magnetic particle spectrum of the unwashed labeled cell sample is calculated (A3-A5) / (A5-A7), which is recorded as K3; In combination with the distribution model and K3, the proportional coefficient of intracellular SPIO and extracellular SPIO in the unwashed labeled cell sample is solved. The proportional coefficient is calculated by: Step S61: Transform the distribution model equation and solve for the proportionality coefficient α: ; Step S62: for the unwashed labeled cell sample to be tested, the odd-order harmonic difference ratio K 3 Bring in , the ratio α of the intracellular SPIO signal in the unwashed labeled cell sample to the total signal U3 is obtained as: ; Step S7: correcting the U3 according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample; In this embodiment, the specific process of correcting the U3 according to the proportional coefficient and obtaining the SPIO signal in the cells of the unwashed labeled cell sample is as follows: Step S71, obtaining a standard unit signal u1 generated by SPIO in the cell, wherein u1 is U1 divided by the total number of cells in the labeled cell sample after washing, wherein the total number of cells in the labeled cell sample is obtained by counting cells when preparing the sample; Step S72, obtaining a standard unit signal u2 generated by extracellular SPIO, wherein u2 is U2 divided by the total number of cells in the sample with ruptured cell membranes, wherein the total number of cells in the sample with ruptured cell membranes is obtained by counting cells when preparing the sample; Step S73, combining the u1 and the u2, calibrating the U3 signal of the unwashed labeled cell sample: ; In the formula, is the signal generated by intracellular SPIO in the unwashed labeled cell sample, and n is the number of cells in the unwashed labeled cell sample.

[0022] In addition, in order to further quickly correct the signal generated by the intracellular SPIO, a rapid correction model for the intracellular SPI signal is constructed. The main difference from the above method is that the former needs to prepare multiple labeled cell samples and perform cell membrane rupture operation on the injected stem cell sample to obtain U2 and K2, while the rapid correction only needs to construct different concentration solutions of labeled particles (PBS dilution) in advance to approximately replace the extracellular signal. In this way, the signal measurement deviation (generated by particles free from cell death) can be calculated by retrieving the K2 curve and U2 curve of the corresponding labeled concentration, and the intracellular signal can be obtained by eliminating the deviation interference. The method is: Step S81, fitting the U1 harmonic curve and K1 curve of the SPIO particle incubated cells R at different magnetic nanoparticle incubation concentrations; Step S82: Based on the characteristics of the voltage response signal U2 of SPIO outside the cell and the voltage response signal U 2-pbs (Approximate) relationships between features, constructing relationships; ; Step S83, fitting the U2-pbs harmonic curve and K2-pbs curve of the SPIO diluted samples of different concentrations, and obtaining the U2 harmonic curve and K2 curve based on the relationship; wherein the U1 harmonic curve, K1 curve, U2 harmonic curve, and K2 curve are obtained by specifically obtaining the point values ​​of each concentration according to the method of steps S1-S4, and then fitting the A3, A5, and A7 harmonic curves of the SPIO particle incubated cells R and the SPIO diluted samples, i.e., the SPIO in the cell and the SPIO outside the cell (see Figure 5 ), and K1 and K 2 Curve (see Figure 6 ); Step S84, according to the SPIO concentration of the incubated cells, respectively search for K1 and K2 on the K1 curve and obtain K1 and K2, and calculate Δ 1 , Δ 2 ; Step S85: The K2, K1, Δ 1 , Δ 2 , the measured cell sample signals U3 and K3 of the unknown SPIO distribution state (specifically obtained according to the method of steps S1-S4) are brought into the proportionality coefficient α to solve the formula, and then the proportionality coefficient of SPIO inside and outside the cell is obtained; ; Step S86, calculating the measurement deviation of the signal based on the relationship and the proportionality coefficient, the measurement deviation of the signal; ; Step S87, based on the measured deviation of the signal, the signal of the cell sample with unknown SPIO distribution state is corrected to obtain the SPIO signal in the cell : .

[0023] This signal correction method uses signal measurement deviation to quickly correct sample signals. It can optimize the image reconstruction model of MPI and other imaging systems, thereby making the signal extraction in cells more accurate and improving the sensitivity of cell tracing. Figure 7 The intraSPIO curve is screened by the model, excluding the SPIO signals that are free outside the cell due to cell death and other reasons, and only retains the spectrum of the response voltage signal generated by SPIO inside the cell.

[0024] Although the various steps in the above embodiment are described in the above-mentioned order, those skilled in the art can understand that in order to achieve the effect of this embodiment, different steps do not have to be executed in such an order. They can be executed simultaneously (in parallel) or in a reverse order. These simple changes are within the scope of protection of the present invention.

[0025] The second embodiment of the present invention is a system for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectrum, which is used to extract and separate intracellular and extracellular SPIO signals, and then obtain the intracellular SPIO signals of unwashed labeled cell samples. The system includes: The MPI signal detection instrument is used to excite and scan the washed labeled cell samples, the samples with ruptured cell membranes, and the unwashed labeled cell samples to obtain response voltage signals U1, U2, and U3; The central processing unit includes a CPU and a GPU; the central processing unit is used to perform Fourier transform on U1 to obtain the magnetic particle spectrum of SPIO in the cell, and then calculate the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K1; Performing Fourier transformation on U2 to obtain the magnetic particle spectrum of extracellular SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K2; Based on the K1 and the K2, constructing a distribution model of SPIO inside and outside the cell; Performing Fourier transformation on the U3 to obtain the magnetic particle spectrum of the unwashed labeled cell sample SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K3; In combination with the distribution model and K3, a proportional coefficient between intracellular SPIO and extracellular SPIO in the unwashed labeled cell sample is solved; The U3 is corrected according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample.

[0026] It should be noted that the quantitative differentiation system for intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy provided in the above embodiment is only illustrated by the division of the above functional modules. In practical applications, the above functions can be assigned to different functional modules as needed, that is, the modules or steps in the embodiments of the present invention can be decomposed or combined. For example, the modules in the above embodiment can be combined into one module, or further divided into multiple sub-modules to complete all or part of the functions described above. The names of the modules and steps involved in the embodiments of the present invention are only for distinguishing the modules or steps, and are not regarded as improper limitations of the present invention.

[0027] An electronic device according to the third embodiment of the present invention comprises: at least one processor; and a memory connected to the at least one processor in communication; wherein the memory stores instructions executable by the processor, and the instructions are used to be executed by the processor to implement the method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy.

[0028] A computer-readable storage medium according to a fourth embodiment of the present invention stores computer instructions, and the computer instructions are used to be executed by the computer to implement the above-mentioned method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy.

[0029] Technicians in the relevant technical field can clearly understand that, for the convenience and brevity of description, the specific working process and related instructions of the electronic device and computer-readable storage medium described above can refer to the corresponding process in the aforementioned method embodiment, and will not be repeated here.

[0030] Those skilled in the art should be able to appreciate that the modules and method steps of each example described in conjunction with the embodiments disclosed herein can be implemented with electronic hardware, computer software, or a combination of the two, and the programs corresponding to the software modules and method steps can be placed in random access memory (RAM), memory, read-only memory (ROM), electrically programmable ROM, electrically erasable programmable ROM, registers, hard disks, removable disks, CD-ROMs, or any other form of storage medium known in the technical field. In order to clearly illustrate the interchangeability of electronic hardware and software, the composition and steps of each example have been generally described in the above description according to the function. Whether these functions are performed in electronic hardware or software depends on the specific application and design constraints of the technical solution. Those skilled in the art can use different methods to implement the described functions for each specific application, but such implementation should not be considered to exceed the scope of the present invention.

[0031] The terms "first", "second", etc. are used to distinguish similar objects rather than to describe or indicate a particular order or sequence.

[0032] The term "comprise" or any other similar term is intended to cover non-exclusive inclusion, such that a process, method, article, or apparatus / device that includes a list of elements includes not only those elements but also other elements not expressly listed, or also includes elements inherent to such process, method, article, or apparatus / device.

[0033] So far, the technical solutions of the present invention have been described in conjunction with the preferred embodiments shown in the accompanying drawings. However, it is easy for those skilled in the art to understand that the protection scope of the present invention is obviously not limited to these specific embodiments. Without departing from the principle of the present invention, those skilled in the art can make equivalent changes or substitutions to the relevant technical features, and the technical solutions after these changes or substitutions will fall within the protection scope of the present invention.

Claims

1. A method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy, which is used to extract and separate intracellular and extracellular SPIO signals, and then obtain the intracellular SPIO signals in unwashed labeled cell samples, characterized in that: The method comprises the following steps: Obtaining a response voltage signal U1 of the washed labeled cell sample under stimulation; Obtaining a response voltage signal U2 of the sample with ruptured cell membrane under excitation; Obtaining a response voltage signal U3 of an unwashed labeled cell sample under stimulation; Perform Fourier transformation on U1 to obtain the magnetic particle spectrum of SPIO in the cell, and then calculate the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K1; Performing Fourier transformation on U2 to obtain the magnetic particle spectrum of extracellular SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K2; Based on the K1 and the K2, constructing a distribution model of SPIO inside and outside the cell; Performing Fourier transformation on the U3 to obtain the magnetic particle spectrum of the unwashed labeled cell sample SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K3; In combination with the distribution model and K3, a proportional coefficient between intracellular SPIO and extracellular SPIO in the unwashed labeled cell sample is solved; The U3 is corrected according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample.

2. The method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to claim 1, characterized in that: The ratio of the difference between the amplitudes of adjacent odd-order harmonics is calculated as: (A3-A5) / (A5-A7), where A3 is the amplitude of the third harmonic, A5 is the amplitude of the fifth harmonic, and A7 is the amplitude of the seventh harmonic.

3. The method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to claim 2, characterized in that: The distribution model of SPIO inside and outside the cell is: ； In the formula, For the labeled cell samples after washing (A5-A7), is the sample with ruptured cell membrane (A5-A7), m is the mixed sample, α is the proportion of the labeled cell sample after washing in the mixed sample, is the proportion of samples with ruptured cell membranes in the mixed samples, It is the ratio of the odd harmonic differences of the mixed samples (A3-A5) / (A5-A7).

4. The method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to claim 3, characterized in that: The proportionality coefficient is calculated as follows: Transform the distribution model equation and solve for the proportionality coefficient α: ； For the unwashed labeled cell sample to be tested, the odd harmonic difference ratio K3 is substituted into , the ratio α of the intracellular SPIO signal in the unwashed labeled cell sample to the total signal U3 is obtained as: 。 5. The method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to claim 1, characterized in that: According to the proportional coefficient, the U3 is corrected to obtain the SPIO signal in the cells of the unwashed labeled cell sample, and the method is: Step S71, obtaining a standard unit signal u1 generated by SPIO in the cell, wherein u1 is U1 divided by the total number of cells in the labeled cell sample after washing; Step S72, obtaining a standard unit signal u2 generated by extracellular SPIO, wherein u2 is U2 divided by the total number of cells in the sample with ruptured cell membranes; Step S73, combining the u1 and the u2, calibrating the U3 signal of the unwashed labeled cell sample: ； In the formula, is the signal generated by intracellular SPIO in the unwashed labeled cell sample, and n is the number of cells in the unwashed labeled cell sample.

6. The method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to claim 5, characterized in that: After obtaining the SPIO signal in the cells of the unwashed labeled cell sample, the method further includes quickly correcting the obtained SPIO signal, the method being: Fitting the U1 harmonic curve and K1 curve of SPIO particle incubated cells R at different magnetic nanoparticle incubation concentrations; According to the voltage response signal U2 of SPIO outside the cell and the voltage response signal U 2-pbs The relationship between features, constructing a relational expression; Fitting the U of SPIO diluted samples with different concentrations 2-pbs Harmonic curve, K 2-pbs Curve, and obtain U2 harmonic curve and K2 curve based on the relationship; According to the SPIO concentration of the incubated cells, K1 and K2 are obtained by searching on the K1 curve and the K2 curve, and Δ1 and Δ2 are calculated based on the corresponding harmonic amplitudes on the U1 harmonic curve and the U2 harmonic curve; Substitute the searched K2, K1, Δ1, Δ2 and the measured cell sample signals U3 and K3 of the unknown SPIO distribution state into the proportional coefficient α solution formula to obtain the proportional coefficient of SPIO inside and outside the cell; Calculating the signal measurement deviation based on the relationship and the proportionality coefficient of the SPIO inside and outside the cell; Based on the measured deviation of the signal, the measured cell sample signal U3 of the unknown SPIO distribution state is corrected to obtain the SPIO signal in the cell .

7. The method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to claim 6, characterized in that: The relationship is: ； The measured deviation of the signal is: ； The intracellular SPIO signal U in : 。 8. A system for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectrum, which is used to extract and separate intracellular and extracellular SPIO signals, and then obtain the intracellular SPIO signals of unwashed labeled cell samples, characterized in that: The system comprises: The MPI signal detection instrument is used to excite and scan the washed labeled cell samples, the samples with ruptured cell membranes, and the unwashed labeled cell samples to obtain response voltage signals U1, U2, and U3; The central processing unit includes a CPU and a GPU; the central processing unit is used to perform Fourier transform on U1 to obtain the magnetic particle spectrum of SPIO in the cell, and then calculate the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K1; Performing Fourier transformation on U2 to obtain the magnetic particle spectrum of extracellular SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd-order harmonics, which is recorded as K2; Based on the K1 and the K2, constructing a distribution model of SPIO inside and outside the cell; Performing Fourier transformation on the U3 to obtain the magnetic particle spectrum of the unwashed labeled cell sample SPIO, and then calculating the ratio of the difference between the amplitudes of adjacent odd harmonics, which is recorded as K3; In combination with the distribution model and K3, a proportional coefficient between intracellular SPIO and extracellular SPIO in the unwashed labeled cell sample is solved; The U3 is corrected according to the proportional coefficient to obtain the SPIO signal in the cells of the unwashed labeled cell sample.

9. An electronic device, characterized in that: include: at least one processor; as well as a memory communicatively connected to at least one of the processors; wherein, The memory stores instructions executable by the processor, and the instructions are used to be executed by the processor to implement the method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy as described in any one of claims 1-7.

10. A computer-readable storage medium storing computer instructions, wherein the computer instructions are used to be executed by the computer to implement the method for quantitatively distinguishing intracellular and extracellular magnetic nanoparticle signals based on magnetic particle spectroscopy according to any one of claims 1 to 7.

Images