Antigen excess monitoring method and monitoring system for specific protein detection

Abstract

The application relates to an antigen excess monitoring method and a monitoring system applied to specific protein detection, and the monitoring method comprises the following steps: firstly, a calibration curve is constructed; secondly, a normal reaction rate and an excess reaction rate are obtained based on the calibration curve; then, a to-be-detected sample reaction process curve of a to-be-detected sample is constructed, and a to-be-detected reaction rate of the to-be-detected sample is obtained through the to-be-detected sample reaction process curve; finally, whether the antigen in the to-be-detected sample is excessive and whether the antigen in the to-be-detected sample is excessive through secondary monitoring are judged according to the to-be-detected reaction rate, the normal reaction rate and the excess reaction rate; the normal reaction rate and the excess reaction rate are determined through the construction of the calibration curve, a double reference standard is formed, and three states of no antigen excess, suspected antigen excess and definite antigen excess can be distinguished more accurately. For the to-be-detected sample in the intermediate interval, a secondary monitoring mechanism is introduced, and the misjudgment probability is further reduced.

Description

Technical Field

[0001] This invention relates to the field of specific protein detection technology, and more particularly to an antigen overload monitoring method and system for specific protein detection. Background Technology

[0002] Specific proteins (also known as special proteins) are a class of proteins with specific physiological or pathological functions, widely present in body fluids such as serum, urine, and cerebrospinal fluid. They participate in various physiological processes such as immune regulation, inflammatory responses, nutritional metabolism, and kidney function. Specific proteins possess highly specific antigenic epitopes; therefore, turbidimetric methods are used to detect them. However, since the detection of specific proteins involves an antigen-antibody reaction, antigen overload is unavoidable, leading to a decrease in the detection signal and false negative results, thus resulting in inaccurate test results. Summary of the Invention

[0003] The purpose of this invention is to provide an antigen overload monitoring method and system for the detection of specific proteins, in order to solve the problem of inaccurate detection results caused by antigen overload in the above-mentioned background art.

[0004] To achieve the above objectives, the technical solution adopted by the present invention is: an antigen overload monitoring method for the detection of specific proteins, comprising the following steps:

[0005] S1. Construct calibration curves based on standards of known concentrations and standard reagents of known concentrations.

[0006] S2. Obtain the normal reaction rate based on the calibration curve. and excess reaction rate ;

[0007] S3. Construct the reaction process curve of the sample to be tested, and obtain the test reaction rate of the sample to be tested from the reaction process curve. ,

[0008] S4. Based on the reaction rate to be detected Normal reaction rate and excess reaction rate Determine whether there is an excess of antigen in the sample to be tested;

[0009] If the reaction rate to be tested Less than normal reaction rate If so, the antigen in the sample to be tested is not excessive;

[0010] If the reaction rate to be tested At normal reaction rate and excess reaction rate Between these steps, secondary monitoring is used to determine whether there is an excessive amount of antigen in the sample to be tested;

[0011] If the reaction rate to be tested Greater than the excess reaction rate If the antigen in the sample to be tested is excessive, then the sample to be tested contains an excess of antigen.

[0012] Beneficial effects: 1) The normal reaction rate is determined by constructing calibration curves. and excess reaction rate This creates a dual reference standard, enabling more precise differentiation between three states: no antigen overdose, suspected overdose, and confirmed overdose. For samples in the intermediate range, a secondary monitoring mechanism is introduced to further reduce the probability of misjudgment, especially avoiding detection deviations caused by ambiguous judgments near the threshold.

[0013] 2) Obtain the reaction rate of the sample to be tested based on the reaction process curve of the sample to be tested. It can dynamically capture changes during the reaction process. Compared with some existing solutions that rely on fixed time point detection, it is more adaptable to the differences in reaction characteristics of different samples. Whether it is a sample with a fast reaction rate or a slow reaction rate, it can obtain accurate rate parameters through curve features, thus improving its adaptability to complex samples.

[0014] 3) By using a tiered judgment logic, conclusions can be drawn directly for samples that are clearly not excessive or clearly excessive, without the need for additional operations; only borderline samples are subject to secondary monitoring, which reduces unnecessary detection steps while ensuring accuracy, thereby improving detection efficiency and reducing time and reagent costs.

[0015] 4) Construct corresponding calibration curves and normal reaction rates for different projects. and excess reaction rate For example, constructing calibration curves for IgG and the normal reaction rate of IgG. and excess reaction rate To construct the calibration curve of IgM and the normal reaction rate of IgM. and excess reaction rate Therefore, this method can be adapted to the detection of multiple items (such as immunoglobulins (IgG, IgM, IgA), complement (C3, C4), C-reactive protein (CRP), urinary microproteins (such as β2-microglobulin) etc.), without the need to develop a separate monitoring system for a single protein, reducing the method development cost and making it suitable for multi-item joint detection scenarios; it is also adapted to dynamic detection processes: real-time monitoring is achieved by collecting signals during the detection process, rather than the traditional endpoint method, which can better capture the dynamic characteristics of antigen excess and improve monitoring sensitivity.

[0016] In a preferred embodiment of the present invention, secondary monitoring includes the following steps:

[0017] S4.1 First, add the secondary monitoring substance to the sample to be tested;

[0018] S4.2, Collect the secondary process scattered light signal value before adding the secondary monitoring substance. ; and the secondary detection scattered light signal value corresponding to a fixed interval after adding the secondary monitoring substance. ;

[0019] S4.3, Based on the scattered light signal value of the secondary process and secondary detection of scattered light signal value Determine whether there is an excess of antigen in the sample to be tested.

[0020] Beneficial effects: 1) By actively adding secondary monitoring substances, the passive waiting for signal changes is transformed into active intervention to observe the reaction trend, thus avoiding the normal reaction rate. and excess reaction rate Excessive sensitivity leading to unnecessary dilution of high-value, normal samples (false positives) also prevents errors due to normal reaction rates. and excess reaction rate The system is designed to detect samples that are too slow to detect early antigen overload (false negatives), thereby reducing the false negative rate and providing a more reliable basis to ensure the accuracy of the final concentration calculation.

[0021] 2) By directly using the secondary process scattered light signal value when the secondary monitoring substance is added. and the second detection of scattered light signal value after a fixed interval. As a basis for judgment, the numerical relationship between the two can clearly indicate whether the antigen is excessive, which is logically clear and the results are intuitive. At the same time, the setting of a fixed interval time ensures the standardization of signal acquisition, makes the secondary monitoring data of different samples comparable, and further improves the accuracy of judgment.

[0022] 3) By adding different types of secondary monitoring substances, it can adapt to different testing scenarios and sample characteristics; whether it is for preliminary screening of antigen overload or retesting of critical samples, it can flexibly meet the testing needs by adjusting the type and interval of the added secondary monitoring substances, which is more versatile than the fixed mode of the existing scheme.

[0023] In a preferred embodiment of the present invention, when the secondary monitoring substance added to the sample to be tested is an antigen,

[0024] If the forward scattered light signal value Less than the value of the scattered light signal detected in the second detection If so, the antigen in the sample to be tested is not excessive;

[0025] Current scattered light signal value Greater than or equal to the value of the secondary detection scattered light signal If so, then there is an excess of antigen in the sample to be tested.

[0026] Beneficial effects: By adding antigens homologous to the target antigen in the sample to be tested, the excess antigen can be determined by comparing the scattered light signal values. This allows for precise focusing on the target reaction and utilizes the principle of specific binding balance in antigen-antibody reactions. The advantages are reflected in the accuracy, specificity, and anti-interference ability of the judgment logic.

[0027] In a preferred embodiment of the present invention, when the secondary monitoring substance added to the sample to be tested is an antibody,

[0028] If the forward scattered light signal value Equal to the value of the scattered light signal detected in the second detection If so, the antigen in the sample to be tested is not excessive;

[0029] Current scattered light signal value Less than the value of the scattered light signal detected in the second detection If so, then there is an excess of antigen in the sample to be tested.

[0030] Beneficial effects: When there is an excess of antigen, the added antibody can bind with the excess antigen to form more complexes, resulting in an enhanced scattered light signal; while when there is no excess antigen, the existing antigen-antibody complexes in the system are relatively stable, and the scattered light signal changes little. This difference in signal change is obvious and can keenly capture the state of antigen quantity. Compared with some existing methods where the signal change is weak and the judgment is ambiguous, this further improves the accuracy of the detection results.

[0031] In a preferred embodiment of the present invention, the calculation steps for the normal reaction rate are as follows:

[0032] S2.1.1 Based on the standards in the calibration curve and the corresponding saturated scattered light signal values ​​of the standards, the concentration calibration point and the signal value calibration point are obtained.

[0033] S2.1.2 Obtain the normal reaction rate from the reaction progress curve of the standard corresponding to the concentration calibration point. .

[0034] Beneficial effects: 1) By combining the standard in the calibration curve with the corresponding saturated scattered light signal value, the concentration calibration point and the signal value calibration point are determined simultaneously, realizing dual anchoring of concentration and signal, providing more comprehensive and stable basic data, and reducing the error caused by fluctuations in single-dimensional data.

[0035] 2) Obtain the normal reaction rate based on the reaction progress curve of the standard corresponding to the concentration calibration point. Instead of relying on theoretical derivation or fixed formula calculations, the reaction process curve can fully reflect the dynamic changes of the standard during the reaction process. The normal reaction rate extracted from it is closer to the actual reaction law, avoiding the problem of rate values ​​deviating from the actual situation due to ignoring the dynamic characteristics of the reaction in existing schemes. This makes the normal reaction rate a more effective reference benchmark and improves the accuracy of the entire antigen overload monitoring system.

[0036] In a preferred embodiment of the present invention, the excess reaction rate is... The calculation steps are as follows:

[0037] S2.2.1 Collect reaction process curves of standard products corresponding to multiple known high-concentration standard products above the concentration calibration point;

[0038] S2.2.2 Based on the reaction progress curves of each high-concentration standard, the basic reaction rate of each standard is obtained. ;

[0039] S2.2.3 Based on multiple basic reaction rates The average value yields the excess reaction rate. .

[0040] Beneficial effects: 1) By using the reaction process curves of multiple standards above the concentration calibration point, these standards are already in the typical state of antigen excess. The reaction rate calculated based on this can more accurately match the reaction law when antigen is in excess in actual detection, avoiding the rate benchmark deviation caused by insufficient representativeness of the standards.

[0041] 2) Calculate the baseline reaction rate for each high-concentration standard individually. The excess reaction rate is then obtained by averaging multiple basic rates. This multi-sample data fusion method can effectively offset individual differences or random errors of a single standard; and reduce the excess reaction rate. It is more stable and more accurate as a critical criterion for judging antigen overdose.

[0042] In a preferred embodiment of the present invention, the normal reaction rate is obtained based on the reaction progress curve of a standard. The steps are as follows:

[0043] S2.1.2.1 Collect two time points in the reaction process curve of the standard corresponding to the concentration calibration point, and the standard process scattered light signal value corresponding to the two time points;

[0044] S2.1.2.2 Obtain the interval time based on the two time nodes, and obtain the difference of the scattered light signal based on the scattered light signal value of the standard sample process corresponding to the two time nodes;

[0045] S2.1.2.3 The normal reaction rate is obtained based on the difference in scattered light signals and the time interval. .

[0046] Beneficial effects: Due to the normal reaction rate The specific time points and the difference in scattered light signal derived from the reaction progress curve of the standard are related to the basic reaction rate. The computational foundation ensures the consistency of both in terms of data dimensions and computational logic; enabling normal reaction rates. and excess reaction rate The interval division is more reasonable, providing a consistent reference standard for judging antigen excess in subsequent samples to be tested, and further improving the accuracy and stability of the entire monitoring system.

[0047] In a preferred embodiment of the present invention, the antigen concentration of the sample to be tested is obtained based on the reaction process curve and calibration curve of the sample to be tested;

[0048] When the antigen in the sample to be tested is not excessive, the saturated scattered light signal value of the sample to be tested is obtained from the reaction process curve of the sample to be tested. Based on the saturated scattered light signal value of the sample to be tested The concentration of the standard corresponding to the calibration curve is the antigen concentration of the sample to be tested;

[0049] When there is an excess of antigen in the sample to be tested, the sample is diluted in a stepwise cycle; the saturated scattered light signal value of the diluted sample is then used as the basis for the determination. The antigen concentration of the sample to be tested is obtained by using the dilution factor and calibration curve.

[0050] Beneficial effects: 1) For samples where the antigen is not in excess, the saturated scattered light signal value of the sample to be tested can be directly utilized. Matching the corresponding concentration in the calibration curve requires no additional processing, reducing errors caused by operational intervention. For samples with excessive antigen, a stepwise cyclic dilution method is used, and the saturated scattered light signal value of the diluted sample is used for analysis. By combining the concentration of the sample to be tested with the dilution factor, detection bias caused by signal saturation or deviation from the linear range due to excessive antigen is effectively avoided, making the detection results of high-concentration samples more accurate.

[0051] 2) The calibration curve is always used as the benchmark. Whether the test sample has no excess antigen or has excess antigen, it is closely related to the standard system of the calibration curve, which ensures the consistency of the test results under the unified standard. At the same time, the step-by-step cyclic dilution operation procedure is standardized and the dilution factor can be accurately recorded, which makes the test process and results have good traceability and facilitates the review and verification of test data in the later stage.

[0052] In a preferred embodiment of the present invention, when there is an excess of antigen in the sample to be tested, the steps for calculating the true concentration of the sample are as follows:

[0053] S5.1 The diluted sample to be tested is obtained by diluting the sample using a stepped cyclic mode;

[0054] S5.2 Acquire the saturated scattered light signal value of the diluted sample to be tested. ,

[0055] S5.2 Based on the saturated scattered light signal value of the diluted sample to be tested The test results of the sample to be tested are obtained by using the signal value calibration points and calibration curves.

[0056] If the saturated scattered light signal value of the sample to be tested is diluted The saturated scattered light signal value of the sample under test is diluted below the signal value calibration point. The antigen concentration in the sample to be tested is obtained by multiplying the concentration corresponding to the calibration curve by the dilution factor.

[0057] If the saturated scattered light signal value of the sample to be tested is diluted If the value is greater than the signal value calibration point, repeat step S5.1.

[0058] Beneficial effects: 1) Stepwise cyclic dilution can adapt to diverse types of samples to be tested (such as serum, urine, cerebrospinal fluid, etc., which may have different concentrations), avoiding missed or false detections due to insufficient or excessive dilution.

[0059] 2) The calibration curve and signal value calibration point are always used as the core reference for concentration calculation. Whether it is initial dilution or cyclic dilution, the correspondence between scattered light signal value and concentration is strictly followed to ensure the comparability of data at different dilution stages. At the same time, the introduction of signal value calibration point provides an objective standard for dilution endpoint, avoids the subjectivity of human judgment, and allows the final concentration result to be derived under a unified scientific framework, further enhancing the repeatability of the results.

[0060] This invention also provides an antigen overload monitoring system for specific protein detection, used to execute the aforementioned antigen overload monitoring method for specific protein detection. The system includes a data acquisition module for acquiring the scattered light signal values ​​of the sample at each time point during the detection process; a time module for ensuring that the interval between acquisitions of two adjacent scattered light signal values ​​is the same; and a processing module for obtaining the scattered light signal difference based on the scattered light signal values ​​at two time points, and simultaneously obtaining the detection reaction rate of the sample based on the scattered light signal difference and the time interval. The judgment module determines the reaction rate to be detected. and the normal reaction rate of storage and excess reaction rate The system determines whether there is an excess of antigen in the sample to be tested, and whether secondary monitoring is used to determine the excess antigen in the sample; the secondary monitoring module is used to determine the reaction rate of the sample to be tested. At normal reaction rate and excess reaction rate The system reconfirms whether the antigen is excessive in the samples to be tested; the dilution module is used to perform stepwise cyclic dilution of the samples with excessive antigen; the output module outputs the antigen concentration in the samples to be tested and the stepwise cyclic dilution process.

[0061] Beneficial effects: 1) Through the acquisition module, time module, processing module, judgment module, secondary monitoring module, dilution module and output module, the entire process from signal acquisition, rate calculation, overload judgment to secondary confirmation, sample dilution and result output is covered; the modules work together to realize the automated connection of detection, judgment and processing, greatly reduce manual intervention and improve detection efficiency and stability;

[0062] 2) Through the linkage design between modules, the processing strategy can be dynamically adjusted according to the actual situation of the sample to be tested: the concentration is directly calculated for samples with no excess antigen, secondary monitoring is initiated for critical samples, and step dilution is automatically triggered for samples with excess antigen. Compared with the fixed process design in the existing technology, it can better adapt to samples with different concentrations and different reaction characteristics, broaden the scope of application of the system, and improve its practicality in the detection of complex clinical samples.

[0063] In addition to the technical problems solved by the present invention, the technical features constituting the technical solutions, and the advantages brought about by the technical features of these technical solutions as described above, other technical problems that the present invention can solve, other technical features contained in the technical solutions, and the advantages brought about by these technical features will be further described in detail with reference to the accompanying drawings. Attached Figure Description

[0064] Figure 1 This is a flowchart of the antigen overload monitoring method of the present invention applied to the detection of specific proteins.

[0065] Figure 2 This is a standard curve diagram of the antigen overload monitoring method for the detection of specific proteins in this invention.

[0066] Figure 3 This is a reaction process curve of the standard corresponding to a standard concentration of 300 mg / L in the antigen overload monitoring method for the detection of specific proteins in this invention.

[0067] Figure 4 This is a graph showing the reaction process of the sample to be tested in the antigen overload monitoring method for the detection of specific proteins according to the present invention.

[0068] Figure 5 This is a graph showing the normal reaction curve of adding antigen twice in the antigen overload monitoring method for the detection of specific proteins according to the present invention.

[0069] Figure 6 This is a graph showing the antigen-antigen excess reaction curve in the antigen excess monitoring method for the detection of specific proteins according to the present invention.

[0070] Figure 7 This is a curve of antibody addition followed by normal reaction in the antigen overload monitoring method for the detection of specific proteins, as described in this invention.

[0071] Figure 8 This is a graph showing the antibody-antigen overload reaction curve in the antigen overload monitoring method for the detection of specific proteins, as described in this invention.

[0072] Figure 9 This is a schematic diagram of the stepwise cyclic dilution method in the antigen overload monitoring method for the detection of specific proteins according to the present invention. Detailed Implementation

[0073] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings.

[0074] Please see Figure 1 As shown, the antigen overload monitoring method for specific protein detection in this embodiment includes the following steps:

[0075] First, several standards of known concentrations (100 mg / L, 200 mg / L, 300 mg / L, 400 mg / L, 500 mg / L, 600 mg / L, 800 mg / L, and 1000 mg / L) were respectively subjected to binding reactions with a standard reagent of known concentration (400 mg / L). The saturated scattering light signal value of each standard (antigen) and standard reagent (antibody) after the binding reaction was obtained. It should be noted that the standard saturated scattered light signal value in this embodiment... This refers to the scattered light signal value corresponding to the maximum and stable number of immune complexes formed when the standard (antigen) is fully bound to the standard reagent (antibody). The saturated scattered light signal value of the standard is used as a reference. Construct calibration curves using the concentrations of standards (e.g.) Figure 2 (As shown).

[0076] According to the calibration curve, when the antigen concentration is 200 mg / L, the corresponding standard saturated scattered light signal value is... The value is 19682, which corresponds to the saturated scattering light signal value of the standard when the antigen concentration is 300 mg / L. The value is 24238, which corresponds to the saturated scattering light signal value of the standard when the antigen concentration is 400 mg / L. The value is 25012; when the antigen concentration is 500 mg / L, the corresponding standard saturated scattering light signal value is... The value is 25712; that is, when the antigen concentration is between 300-500 mg / L, the increase in the scattered light signal value slows down, indicating that it is approaching the "equivalence band" of antigen-antibody binding. Therefore, 300 mg / L is used as the concentration calibration point, and the standard saturated scattered light signal value corresponding to 300 mg / L is... As a calibration point for signal values.

[0077] During the combination reaction of the standard and standard reagent at each concentration, the starting point of the combination was taken as the starting point, and the scattered light signal value was continuously read at fixed time intervals (t). A standard reaction progress curve was constructed with time as the X-axis and the scattered light signal value as the Y-axis (e.g., ...). Figure 3 (As shown).

[0078] The normal reaction rate was obtained from the reaction progress curve of the standard sample corresponding to the concentration calibration point. ; to obtain the normal reaction rate The method is as follows:

[0079] Please see Figure 3 As shown, two time points were first collected within the reaction progress curve of the standard ( , ) and the standard process scattered light signal values ​​at the two time points ( , It should be noted that the time point The optimal time point for the binding reaction to be carried out using standard (antigen) and standard reagent (antibody) is... Time point Preferred time point Adjacent time points. Based on the scattered light signal values ​​of the two standard samples ( , The difference in scattered light signals was obtained. The interval between the two time points is obtained. The normal reaction rate is obtained based on the time interval and the difference in scattered light signals. Normal reaction rate The calculation formula is as follows:

[0080] = =( - ) / ( - );

[0081] In this embodiment, a standard with a concentration of 300 mg / L is used as the concentration calibration point, and the corresponding normal reaction rate is obtained based on the standard reaction process curve corresponding to the standard with a concentration of 300 mg / L. For example, if the starting point of the reaction process curve of the standard is taken as t1, then t1=15, and the time point t2=20, then S2=14656. =100, therefore according to the normal reaction rate The calculation formula can be obtained =2911.2.

[0082] The basic reaction rate is obtained by selecting the reaction process curves of multiple standards with concentrations higher than the concentration calibration point. Finally, based on multiple fundamental reaction rates Obtain the excess reaction rate This embodiment uses multiple basic reaction rates. The arithmetic mean yields the excess reaction rate. In other embodiments, the excess reaction rate can be obtained by other means (such as weighted average, geometric average). It should be noted that the basic reaction rate Calculation method and normal reaction rate The calculation method is the same. It will not be repeated here.

[0083] For example, the basic reaction rate can be obtained by selecting the reaction process curve of a standard with a concentration of 400 mg / L. The basic reaction rate was obtained from the reaction progress curve of the standard with a concentration of 600 mg / L. The basic reaction rate was obtained from the reaction progress curve of the standard with a concentration of 800 mg / L. The basic reaction rate was obtained from the reaction progress curve of the standard with a concentration of 1000 mg / L. Therefore, the excess reaction rate = ( + + + ) / 4.

[0084] Then, the sample to be tested is tested. First, a reaction curve of the sample to be tested and the test reagent is established (e.g., Figure 4 As shown in the figure, the reaction rate to be tested is obtained from the reaction process curve of the sample to be tested. The method for constructing the reaction process curve of the sample to be tested is the same as that for constructing the reaction process curve of the standard sample, and the reaction rate to be tested is... And the above-mentioned normal reaction rate The calculation method is the same, and will not be repeated here. Simultaneously, the time points in the reaction process curve of the sample to be tested and the above-mentioned normal reaction rate are collected. The time points are the same.

[0085] Based on the reaction rate to be detected Normal reaction rate and excess reaction rate Determine whether the antigen in the sample to be tested is in excess; if the reaction rate of the sample to be tested is... Less than normal reaction rate The sample concentration was determined to be within the normal linear measurement range, indicating that the antigen in the sample was not excessive. The saturated scattered light signal value of the sample was determined based on the reaction process curve of the sample. The concentration of the standard corresponding to the calibration curve is the concentration of the antigen in the sample to be tested.

[0086] If the reaction rate to be tested At normal reaction rate and excess reaction rate Between these values ​​is the ambiguous zone / to be confirmed. This situation is in a gray area of ​​decision-making. It may be a high-value sample with a concentration close to the upper limit of the linear range, or it may be a sample that has just entered the antigen excess zone. Therefore, secondary monitoring is performed to determine whether the antigen in the sample to be tested is excessive.

[0087] Please see Figure 5-6 As shown, during secondary monitoring, the antigen is first added to the sample to be tested, and the forward scattering light signal value before the antigen is added is recorded. .

[0088] Then, after a fixed interval, the secondary detection scattered light signal values ​​corresponding to adjacent time points are collected. Finally, based on the forward-scattered light signal value and secondary detection of scattered light signal value Determine whether there is an excess of antigen in the sample to be tested;

[0089] If the forward scattered light signal value Less than the value of the scattered light signal detected in the second detection If the antigen in the sample to be tested is not excessive, then the antigen content is not excessive. Figure 5 (as shown)

[0090] Current scattered light signal value Greater than or equal to the value of the secondary detection scattered light signal If the antigen in the sample to be tested is excessive (e.g., Figure 6 (As shown).

[0091] If the reaction rate to be tested Greater than the excess reaction rate This indicates an extremely rapid reaction rate, far exceeding the upper limit of the normal linear range, a typical characteristic of antigen overload. If there is antigen overload in the sample to be tested, a stepwise cyclic dilution (e.g., ...) should be performed on the current sample. Figure 9 (as shown in the figure) and obtain the true concentration of the sample to be tested.

[0092] First, a stepped cyclic method is used to dilute the sample to be tested to obtain a diluted sample to be tested.

[0093] Then, the saturated scattered light signal value of the diluted sample to be tested was collected. ;

[0094] Finally, based on the saturated scattered light signal value of the diluted sample to be tested... The test results of the sample to be tested are obtained by using the signal value calibration points and calibration curves.

[0095] If the saturated scattered light signal value of the sample to be tested is diluted The saturated scattered light signal value of the sample under test is diluted below the signal value calibration point. The antigen concentration in the sample to be tested is obtained by multiplying the concentration corresponding to the calibration curve by the dilution factor.

[0096] If the saturated scattered light signal value of the sample to be tested is diluted If the value is greater than the signal calibration point, a step-by-step cyclic mode is used to dilute the sample to be tested.

[0097] For example, a sample to be tested is diluted one order to obtain a diluted sample (a fixed dilution factor, such as 8 times); the dilution process of the sample is monitored and a reaction process curve of the diluted sample is constructed; based on the reaction process curve of the diluted sample, the saturated scattered light signal value of the diluted sample is obtained. If the saturated scattered light signal value of the sample to be tested is diluted The saturated scattered light signal value of the sample under test is diluted below the signal value calibration point. The antigen concentration in the sample to be tested is obtained by multiplying the concentration corresponding to the calibration curve by the dilution factor; then the true concentration = concentration * first-order dilution factor (8), if the saturated scattered light signal value of the diluted sample to be tested is... If the value is greater than the calibration point, then proceed with a second-order dilution (a second dilution at a fixed factor, such as 5 times, based on the first-order dilution). The second-order diluted sample corresponds to the saturated scattered light signal value of the diluted sample. If the signal value is less than the calibration point, the true concentration = concentration * first-order dilution factor (8) * second-order dilution factor (5). For example, the saturated scattered light signal value of the diluted sample after second-order dilution. If the concentration still exceeds the range of the standard curve, initiate a third-order dilution (dilution again based on the first and second orders) until the results are obtained.

[0098] Please see Figure 7-8 As shown, the difference between Example 2 and Example 1 is that, during the secondary monitoring, an antibody is added to the sample to be tested, and the forward scattering light signal value before the addition of the antibody is recorded. ;

[0099] Then, after a fixed interval, the secondary detection scattered light signal values ​​corresponding to adjacent time points are collected. Finally, based on the forward-scattered light signal value and secondary detection of scattered light signal value Determine whether there is an excess of antigen in the sample to be tested.

[0100] If the forward scattered light signal value Equal to the value of the scattered light signal detected in the second detection If the antigen in the sample to be tested is not excessive (e.g., Figure 7 (as shown)

[0101] Current scattered light signal value Less than the value of the scattered light signal detected in the second detection If the antigen in the sample to be tested is excessive (e.g., Figure 8 (As shown).

[0102] This invention also provides an antigen overload monitoring system for specific protein detection, used to execute the aforementioned antigen overload monitoring method for specific protein detection. The system includes a data acquisition module for acquiring the scattered light signal values ​​of the sample at each time point during the detection process; a time module for ensuring that the interval between acquisitions of two adjacent scattered light signal values ​​is the same; and a processing module for obtaining the scattered light signal difference based on the scattered light signal values ​​at two time points, and simultaneously obtaining the detection reaction rate of the sample based on the scattered light signal difference and the time interval. The judgment module determines the reaction rate to be detected. and the normal reaction rate of storage and excess reaction rate The secondary monitoring module is used to determine whether there is an excess of antigen in the sample to be tested, and whether secondary monitoring is used to determine the excess of antigen in the sample to be tested; the secondary monitoring module is used to determine the reaction rate of the sample to be tested. At normal reaction rate and excess reaction rate The system reconfirms whether the antigen is excessive in the samples to be tested; the dilution module is used to perform stepwise cyclic dilution of the samples with excessive antigen; the output module outputs the antigen concentration in the samples to be tested and the stepwise cyclic dilution process.

[0103] It should be noted that all time units in the specification and accompanying drawings of this application are seconds.

[0104] If the embodiments of the present invention involve directional indicators (such as up, down, left, right, front, back, etc.), the directional indicators are only used to explain the relative positional relationship and movement of the components in a certain specific posture (as shown in the figure). If the specific posture changes, the directional indicators will also change accordingly.

[0105] The above embodiments are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A method for monitoring antigen overload in the detection of specific proteins, characterized in that, Includes the following steps: S1. Construct a calibration curve based on standards of known concentrations. S2. Obtain the normal reaction rate based on the calibration curve. and excess reaction rate The concentration calibration point is obtained from the calibration curve. The normal reaction rate is obtained from the time interval between any two time points in the reaction progress curve of the standard corresponding to the concentration calibration point and the difference in the scattered light signal between the two time points. The excess reaction rate is obtained by considering the time interval between any two time points in the reaction progress curves of standards above the concentration calibration points and the difference in the scattered light signals between the two time points. The point where the increase in the value of the scattered light signal slows down and approaches the equivalence band of antigen-antibody binding is used as the concentration calibration point; S3. Construct the reaction process curve of the sample to be tested, and obtain the reaction rate of the sample to be tested through the reaction process curve. S4. Based on the reaction rate to be detected Normal reaction rate and excess reaction rate Determine whether there is an excess of antigen in the sample to be tested; If the reaction rate to be tested Less than normal reaction rate If so, the antigen in the sample to be tested is not excessive; If the reaction rate to be tested At normal reaction rate and excess reaction rate Between these steps, secondary monitoring is used to determine whether there is an excessive amount of antigen in the sample to be tested; If the reaction rate to be tested is greater than the excess reaction rate If the antigen in the sample to be tested is excessive, then the sample to be tested contains an excess of antigen.

2. The antigen overload monitoring method for specific protein detection according to claim 1, characterized in that, Secondary monitoring includes the following steps: S4.1 First, add the secondary monitoring substance to the sample to be tested; S4.2, Collect the forward-scattered light signal value before adding the secondary monitoring substance. ; and the secondary detection scattered light signal value corresponding to a fixed interval after the addition of the secondary monitoring substance. ; S4.3, Based on the forward-scattered light signal value The value of the scattered light signal from the secondary detection is used to determine whether there is an excess of antigen in the sample to be tested.

3. The antigen overload monitoring method for specific protein detection according to claim 2, characterized in that: When the secondary monitoring substance added to the sample to be tested is an antigen, If the forward scattered light signal value Less than the value of the scattered light signal detected in the second detection If so, the antigen in the sample to be tested is not excessive; Current scattered light signal value Greater than or equal to the value of the secondary detection scattered light signal If so, then there is an excess of antigen in the sample to be tested.

4. The antigen overload monitoring method for specific protein detection according to claim 2, characterized in that: When the secondary monitoring substance added to the sample to be tested is an antibody, If the forward scattered light signal value Equal to the value of the scattered light signal detected in the second detection If so, the antigen in the sample to be tested is not excessive; Current scattered light signal value Less than the value of the scattered light signal detected in the second detection If the antigen content in the sample to be tested is excessive, then the sample to be tested will contain an excessive amount of antigen.

5. The antigen overload monitoring method for specific protein detection according to claim 1, characterized in that: The steps for calculating the normal reaction rate are as follows: S2.1.1 Based on the standards in the calibration curve and the corresponding saturated scattered light signal values ​​of the standards, the concentration calibration point and the signal value calibration point are obtained. S2.1.2 Obtain the normal reaction rate from the reaction progress curve of the standard corresponding to the concentration calibration point. .

6. The antigen overload monitoring method for specific protein detection according to claim 5, characterized in that: Excess reaction rate The calculation steps are as follows: S2.2.1 Collect reaction process curves of standard products corresponding to multiple known high-concentration standard products above the concentration calibration point; S2.2.2 Based on the reaction progress curves of each known high-concentration standard, the basic reaction rate of each standard is obtained. ; S2.2.3 Based on multiple basic reaction rates The average value yields the excess reaction rate. .

7. The antigen overload monitoring method for specific protein detection according to claim 5, characterized in that: The normal reaction rate was obtained from the reaction progress curve of the standard sample. The steps are as follows: S2.1.2.1 Collect two time points in the reaction process curve of the standard corresponding to the concentration calibration point, and the standard process scattered light signal value corresponding to the two time points; S2.1.2.2 Obtain the interval time based on the two time nodes, and obtain the difference of the scattered light signal based on the scattered light signal value of the standard sample process corresponding to the two time nodes; S2.1.2.3 The normal reaction rate is obtained based on the difference in scattered light signals and the time interval. .

8. The antigen overload monitoring method for specific protein detection according to claim 5, characterized in that: The antigen concentration of the sample to be tested was obtained based on the reaction process curve and calibration curve. When the antigen in the sample to be tested is not excessive, the saturated scattered light signal value of the sample to be tested is obtained from the reaction process curve of the sample to be tested. Based on the saturated scattered light signal value of the sample to be tested The concentration of the standard corresponding to the calibration curve is the antigen concentration of the sample to be tested; When there is an excess of antigen in the sample to be tested, the sample is diluted in a stepwise cycle; the saturated scattered light signal value of the diluted sample is then used as the basis for the determination. The antigen concentration of the sample to be tested is obtained by using the dilution factor and calibration curve.

9. The antigen overload monitoring method for specific protein detection according to claim 8, characterized in that: When there is an excess of antigen in the sample to be tested, the steps for calculating the true concentration of the sample are as follows. S5.1 The diluted sample to be tested is obtained by diluting the sample using a stepped cyclic mode; S5.2 Acquire the saturated scattered light signal value of the diluted sample to be tested. , S5.2 Based on the saturated scattered light signal value of the diluted sample to be tested The test results of the sample to be tested are obtained by using the signal value calibration points and calibration curves. If the saturated scattered light signal value of the sample to be tested is diluted The saturated scattered light signal value of the sample under test is diluted below the signal value calibration point. The antigen concentration in the sample to be tested is obtained by multiplying the concentration corresponding to the calibration curve by the dilution factor. If the saturated scattered light signal value of the sample to be tested is diluted If the value is greater than the signal value calibration point, repeat step S5.

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10. An antigen overload monitoring system for the detection of a specific protein, used to perform the antigen overload monitoring method for the detection of a specific protein as described in any one of claims 1-9, characterized in that: It includes a data acquisition module, used to acquire the scattered light signal values ​​of the sample under test at each time point during the testing process. The time module is used to ensure that the interval between the acquisition of two adjacent scattered light signal values ​​is the same; The processing module calculates the difference between the scattered light signals at two time points, and simultaneously obtains the reaction rate of the sample to be tested based on the difference between the scattered light signals and the time interval. ; The judgment module determines the reaction rate to be detected. and the normal reaction rate of storage and excess reaction rate To determine whether there is an excess of antigen in the sample to be tested, and whether to use secondary monitoring to determine whether there is an excess of antigen in the sample to be tested; The secondary monitoring module is used to monitor the reaction rate of the target. At normal reaction rate and excess reaction rate The samples to be tested were reconfirmed to determine whether there was an excess of antigen. The dilution module is used for stepwise cyclic dilution of test samples with excessively high antigen levels. The output module outputs the antigen concentration in the sample to be tested and the stepwise cyclic dilution process.

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