CN110261592B

CN110261592B - Qualitative and quantitative determination method for analyte

Info

Publication number: CN110261592B
Application number: CN201910606520.XA
Authority: CN
Inventors: 孙少民
Original assignee: Hangzhou Bufeng Technology Co ltd
Current assignee: Hangzhou Bufeng Technology Co ltd
Priority date: 2019-07-05
Filing date: 2019-07-05
Publication date: 2023-06-02
Anticipated expiration: 2039-07-05
Also published as: CN110261592A

Abstract

The invention provides a qualitative and quantitative determination method of an analyte, which comprises the steps of automatic sample adding, incubation, transferring, cleaning and detection, wherein the incubation and the cleaning are respectively carried out on an incubation module and a cleaning module, a group of solid phase units in different reaction phases start and finish incubation on the incubation module and start and finish cleaning on the cleaning module at the same time; according to the invention, based on the fact that each step of reaction of the analyte is required to be incubated and cleaned, a group of solid phase units in each step of reaction stage are synchronously incubated and cleaned through automatic transfer, on one hand, single sample sequential detection is realized, the processing conditions of each sample are consistent, and the detection result is more accurate; on the other hand, each group of solid phase units at different stages can be simultaneously incubated and cleaned, so that the detection efficiency is greatly improved.

Description

Qualitative and quantitative determination method for analyte

Technical Field

The invention relates to the technical field of immunoassay, in particular to a qualitative and quantitative determination method for an analyte.

Background

Phenomena of specific ligand-specific spontaneous binding, such as immunoassay of antigen-antibody specific reaction and nucleic acid analysis of nucleic acid base complementary pairing, are widely used in various fields. Among all the analysis methods, the immunoassay type is the most complicated. But in general there may be four dimensional classifications, namely single or multiple, heterogeneous or homogeneous, simultaneous or non-simultaneous treatments, separation and no separation.

Planar or membrane-based microarrays (solid-phase biochips) and flow cytometry-based liquid-phase chips with encoded microspheres as carriers can detect multiple analytes simultaneously, divided in a first dimension, while other methods are generally capable of detecting only one analyte.

Planar or membrane-based microarrays (solid-phase biochips) represent the fundamental feature of heterogeneous phase immunoassays according to a second dimension division, ELISA, immunochromatography. Based on the above technology, other technologies, such as microarray solid phase immunoassay, plate chemiluminescence, time resolution, fluorescence immunochromatography, etc., are also derived. The liquid phase chip based on flow cytometry and taking the encoded microsphere as a carrier represents the basic characteristics of homogeneous immune reaction.

In a third dimension, since immune responses generally involve multiple steps of antigen-antibody reactions and separations, two strategies are basically employed to meet realistic demands. The simultaneous treatment is to treat a certain step, such as ELISA and similar plate chemiluminescence, in batch at the same time. The advantage of simultaneous treatment is large throughput and low reagent cost. Is particularly suitable for large-scale detection tasks.

According to the fourth dimension division, the liquid phase chip based on the flow cytometry and taking the coded microspheres as a carrier and the photo-excitation chemiluminescence can theoretically realize the analyte determination under the condition of non-separation. Other methods all require a separation step.

How to accurately, efficiently, flexibly, with low cost, and continuously and stably obtain analysis results is the aim pursued by the invention. Among the techniques described above, there are several techniques for achieving this goal:

ELISA or plate chemiluminescence. The technology is based on the design of microwell plates, where an antigen or antibody associated with an analyte is immobilized within a microwell for use with an antigen or antibody of a liquid to analyze an analyte in a liquid sample. When in use, a user uses the micropore plate frame as a supporting body, micropores or whole plates or single plates are combined, and the detection results of the same analyte of a plurality of samples are finally obtained through a plurality of steps of incubation (antigen-antibody reaction) and separation (washing). The separation device used therein uses a vacuum pump to reject waste liquid in the wells, and uses peristaltic or priming pumps and multiple channels to clean the wells. Typically, the spray head and tip are multi-channel designs, with a maximum of 96 channels and a minimum of 8 channels. In this method, additional microwells are typically arranged to measure the sample while simultaneously measuring the calibrator. For qualitative products, it is often necessary to determine a negative control, a positive control and a cutoff control at the same time as the sample, and determine the qualitative result of the sample by comparison with the controls; for quantitative products, it is often necessary to measure a series of calibrators, such as analyte standard substances having a concentration from 0 to a certain value, simultaneously with the measurement of the sample, and calculate the concentration of the analyte in the sample by means of a calibration curve drawn with the concentration of the calibrator and the signal of the calibrator.

In contrast to the above techniques, microarray ELISA methods, such as those available from Quansysbio, hua Daji Biai, ming Yuan Dukang, etc., have been developed. A high precision spotting machine, such as a piezoelectric spotting device from Bio Dot corporation, is used to spot a plurality of antigens or antibodies associated with different analytes onto the bottom of a microplate, which may be polystyrene or nitrocellulose membrane. The reaction section was completed in a manner consistent with ELISA, and finally, the method was different from the method of measuring optical density by ELISA, and the optical signal intensities of points at different positions of the bottom of the microwell exposed for a long time were recorded using CCD to obtain qualitative or quantitative results.

However, this approach does not address the detection problem of a single sample. This approach relies on the fact that a set of calibrators must be measured simultaneously with the measurement of the sample to calculate the concentration of the analyte in the current sample.

The Chinese patent (Suzhou triple living things) solves the problems, and adopts the inserted hard glass as a carrier, and the detection is completed by moving the hard chip. However, the chip has a plurality of problems, including a corresponding flushing and drying mode designed based on the glass hard chip, and the chip has a complex structure and is not beneficial to uninterrupted cleaning.

Separation is an essential step in the above two methods, and the strategy for the implementation of this process is multi-channel washing as described above. In practical applications, clogging of the suction and disposal channels is a frequently encountered problem due to the complex composition of the sample. To solve this problem, plate washing techniques based on the centrifugal principle have been developed, such as chinese patent CN202683525, blue Cat Bio company's Bluewahser; and ejecting the inverted microplate using an ejection head. The method they use is characterized by "simultaneous processing", i.e. washing either in whole plate units or in whole strip units. This washing method has the following problems: 1. the structure is complex and energy is consumed; 2. the rapid washing treatment of a single sample cannot be achieved; 3. the liquid path is in direct contact with the sample, so that frequent maintenance is needed, and cross contamination is avoided.

All the methods described above, which have the feature of "simultaneous processing", do not actually "simultaneously process" the biological reaction being performed in practical applications. Because the samples to be analyzed are added one by one, the time required to add the samples can be ignored if the number of samples is small, but if the number of samples is increased, such as 100 samples, the time interval from the first sample to the last sample will be more than 10 minutes. This approach is especially relevant for those test methods based on the competition principle. The meaning of "simultaneous processing" is only that it leaves the user labor to be reduced, which is detrimental to accurate measurements and increases the deviation of the measurement results as the sample size increases.

Therefore, for the accuracy of the detection, it is most desirable that the conditions of each step from sample addition to final result measurement and the transition time between steps be kept identical or the difference be negligible. However, multi-step wash separations and tens of hundred sample assays will increase the time of detection, severely impacting the practical acceptance of this ideal situation. Increasing the reaction rate and increasing the number of cleaning elements can effectively improve the practical acceptance.

The current homogeneous immune reaction using magnetic particles as carrier effectively solves the problem. Because of the rapid homogeneous reaction, the single-step immune reaction can be shortened to 10 minutes, and the rapid result can be achieved by adding cleaning elements. Such as the method disclosed in patent CN 201310300162.2. The first test result may be obtained at 18 minutes, followed by one test result every 20 seconds. However, this method can only achieve one result at a time. Multiple results cannot be obtained at one time.

Often, multiple outcomes are more helpful in disease judgment. For example, the combined use of several tumor markers can determine the early state of some tumors more accurately or recognize earlier, and for example, the detection of multiple subtypes of HPV can prevent the detection of pathogens missing cervical cancer, etc.

Liquid phase chip technology using flow cytometry appears to satisfactorily address the objective of simultaneous determination of multiple analytes at a time. However, this technique relies on two fluorescent materials as the labeled microsphere structure. The surface of the microsphere is also provided with fluorescent substances, the fluorescent substances in nature and artificial synthesis have hydrophobic property, the substances spontaneously aggregate in aqueous solution, the aggregation of the hydrophobic effect has an accumulation effect, and therefore, the aggregation of different degrees is caused by long-time storage or environmental induction (such as evaporation of the surface of a reagent liquid), so that the technology needs to be frequently corrected and calibrated in practical application.

In addition, the hydrophobic nature of the surface of such spheres is disadvantageous for measurement of samples with high lipid concentrations and is limited in clinical applications. Because all reagents are in a liquid state, the degree of freedom of the reagents is increased, the risk of variation is increased, and in order to efficiently and sensitively detect single microspheres, hundreds of microspheres must be passed through a capillary tube in a detector at a high speed in a single microsphere mode, so that the physical use frequency of the capillary tube is too high, and a tube blockage phenomenon is easy to occur. The risk of failure after long-term operation of the method is increased.

From the viewpoint of information flow, the liquid phase chip is firstly produced by one microsphere, then mixed together, reacted with a sample, and then different microspheres are arranged into a straight line by using external force, and analyzed one by using laser excitation and a microscope in a short distance. This process of analysis from ordered to unordered to ordered is inherently very complex and unsuitable as a large scale and stable analysis method.

The steps of the solid phase chip which are carried out step by step are clear, and the steps are not mutually interfered by each other through stable and rapid separation, so that various problems of the other methods in practical application can be avoided. The invention aims to provide a qualitative and quantitative determination method, which can sequentially determine samples without interruption and can obtain a plurality of test results in each determination.

Disclosure of Invention

In order to solve the problems, the invention provides a qualitative and quantitative determination method of an analyte, which comprises automatic sample adding, incubation, cleaning and detection, wherein the incubation and the cleaning are respectively carried out on an incubation module and a cleaning module, and a group of solid phase units in different reaction stages start and finish incubation on the incubation module and start and finish cleaning on the cleaning module simultaneously.

In some preferred modes, the incubation module comprises at least one group of solid phase unit accommodation sites arranged in a straight line, each group of solid phase unit accommodation sites comprises a new sample loading site and a sample to be tested at two ends of a split row, and one group of solid phase units in each reaction stage are placed in the corresponding accommodation sites to perform incubation reaction. The solid phase units of each reaction stage are placed in corresponding solid phase unit accommodating positions, so that the separation and the distinction of the solid phase units of different reaction stages are facilitated, and the multi-step reaction of the solid phase units is facilitated.

In some preferred embodiments, each set of solid phase unit receptacles of the incubation module comprises N1 solid phase unit receptacles, where n1=3n or 2n, N fresh sample receptacles, and N test receptacles are arranged across the array. For the three-step method or the two-step method, the containing positions of the solid phase units with different numbers can be arranged, and the number of the incubation reaction steps corresponds to the number of the containing positions.

In some preferred forms, n=1 or 2 or 3. Considering the influence of the sample adding time on accurate measurement, the number of the solid phase units added with samples at each time is not excessive, and the detection efficiency is further improved on the basis of ensuring the accuracy of detection results by newly adding 1 or 2 or 3 solid phase units at each time.

In some preferred modes, after a group of solid phase units are washed, the solid phase units on the sample position to be detected of the original incubation module are detected, the rest solid phase units are transported to the incubation module and are shifted from the new sample adding position to the direction of the sample position to be detected on the incubation module, and meanwhile, the new solid phase units after sample adding are supplemented to the new sample adding position, and the incubation is performed again. After the incubation and cleaning of the solid phase units on the sample position to be tested of the incubation module are finished, the rest solid phase units need to enter the next incubation and cleaning process because the incubation and cleaning processes are finished once, so that when the solid phase units are transferred back to the incubation module, the corresponding containing positions are changed, namely, the solid phase units on the original new sample position are moved into the containing positions of the solid phase units after the new sample position, the new sample position is used for containing the new solid phase units, and the solid phase units for the last incubation are moved into the sample position to be tested. Thus, the solid phase units in different reaction stages can be simultaneously subjected to incubation and cleaning operations, and single solid phase units are sequentially subjected to detection.

In some preferred embodiments, after the last set of solid phase units has been washed and incubation is resumed, the next set of solid phase units is subjected to a washing, re-incubation step, until the last set, and sequentially cycled. And the number of the solid phase unit accommodation sites of the incubation module is N < 2 > =T/T, wherein T is the interval time from adding the solid phase unit into the solid phase unit accommodation site of the incubation module to adding the solid phase unit accommodation site of the incubation module again, and T is the interval time from moving one group of solid phase units into the solid phase unit accommodation site of the incubation module to moving the next group of solid phase units into the solid phase unit accommodation site of the incubation module. Because the incubation reaction time is longer than the transfer time of the solid phase units between the incubation module and the cleaning time in the cleaning module, by setting the T/T group of solid phase units, the next group of solid phase units complete the incubation reaction after the last group of solid phase units complete cleaning and are transferred to the incubation module again, and the cleaning operation can be performed until the last group of solid phase units are circulated in sequence. Because the groups are not connected with each other in a waiting way or in a very short time, the time of the whole detection process is saved, and the detection efficiency is greatly improved.

In some preferred embodiments, the incubation module comprises a long plate-shaped incubation plate on which each set of solid phase unit accommodation sites is arranged in parallel along the length direction. The long plate-shaped incubation plate has a simple structure, and is convenient for each group of solid phase units to incubate and clean in sequence.

In some preferred forms, the incubation module comprises a disc-shaped incubation disc on which the sets of solid phase unit accommodation sites are arranged in a circumferential direction, and the line between the midpoint of the line between each set of solid phase unit accommodation sites and the center of the circle is perpendicular to the line between each set of solid phase unit accommodation sites or the extension line of the line between each set of solid phase unit accommodation sites passes through the center of the circle.

In some preferred forms, the bottom of the disc-shaped incubation plate is provided with a rotating device, the incubation plate is further provided with a cover body, the cover body is provided with an opening for exposing one group of solid phase units, after one group of solid phase units is washed and incubation is started again, the rotating device drives the incubation plate to rotate, the cover body is not moved, the opening of the cover body exposes the next group of solid phase units, and the washing and incubation steps are carried out again. The disc-shaped incubation plate is adopted and the rotating device is arranged, so that the cavity space between the incubation plate and the cover body can be reduced, and the stable incubation reaction environment can be provided.

In some preferred forms, the method further comprises an automatic transfer step, the automatic transfer being accomplished by a transfer module comprising a fresh sample transfer module, a wash transfer module, a test sample transfer module, a first gripper, a second gripper, a third gripper, the fresh sample transfer module comprising N3 fresh sample addition sites of solid phase units, wherein n3=n; the to-be-detected sample transfer module comprises N4 to-be-detected sample positions of the solid phase unit, wherein N4=n. The transfer module is used for transferring the solid phase unit among the solid phase unit storage module, the incubation module, the cleaning module and the measurement module, and under the combined action of the new sample loading transfer module, the cleaning transfer module, the sample transfer module to be tested, the first gripper, the second gripper and the third gripper, the automatic transfer of the solid phase unit among the modules is realized, and the detection efficiency is improved.

In some preferred embodiments, the washing and transferring module comprises a set of N5 solid phase unit accommodation sites aligned in a line, where n5=n1, that is, the number of solid phase unit accommodation sites of the washing and transferring module is the same as the number of solid phase unit accommodation sites of the incubation module, and the solid phase unit accommodation sites include N solid phase unit sample sites at one end.

The new sample adding and transferring module and the cleaning and transferring module are positioned on the same straight line, and reciprocate along the straight line, and when the new sample adding and transferring module moves to the position I, the first gripper moves along the straight line to transfer the solid phase unit from the solid phase unit storage module to the new sample adding position of the new sample adding and transferring module; when the sample moves to the position II, sample adding is carried out; and when the sample is moved to the position III, the new sample adding position of the new sample adding transfer module is aligned with the new sample adding position of the incubation module in parallel.

The cleaning and transferring module moves in a linear reciprocating mode, when the cleaning and transferring module moves to a first position, a sample position to be tested of the cleaning and transferring module is aligned with a sample position to be tested of the incubation module in parallel, and a group of solid phase units are transferred onto the cleaning and transferring module from the incubation module in a linear mode through second handles; when the solid phase unit of the cleaning and transferring module finishes adding the washing liquid and moves to the second position after cleaning, the sample position to be tested of the cleaning and transferring module is aligned with the sample position to be tested of the sample transferring module in parallel, and the solid phase unit on the sample position to be tested of the cleaning and transferring module is transferred to the sample position to be tested of the sample transferring module along the linear motion, and the solid phase unit to be tested is transferred to the measuring module for detection by the sample transferring module; when the cleaning and transferring module moves to the third position, the cleaning and transferring module is closely adjacent to the new sample adding and transferring module located at the position III, and the second gripper transfers the new sample adding and transferring module and the solid phase unit on the cleaning and transferring module to the incubation module along the linear motion.

Through the cooperation of new application of sample transport module, washing transport module, the movement of the sample transport module that awaits measuring and position and the transfer effect of first tongs, second tongs, third tongs, realized the solid phase unit and by the aversion of new application of sample position to the sample position that awaits measuring at incubating the module, realized that single solid phase unit is full-automatic detects in order, improved detection efficiency on the basis of improving the detection accuracy.

In some preferred embodiments, the washing transfer module comprises a set of N5 solid phase unit receptacles in a linear arrangement, where n5=n1+n, the solid phase unit receptacles comprising N solid phase unit fresh sample addition locations and N solid phase unit test samples at both ends.

The new sample adding and transferring module moves in a reciprocating manner along a straight line, and when the new sample adding and transferring module moves to the position I, the first gripper moves along the straight line, and the solid phase unit is transferred to a new sample adding position of the new sample adding and transferring module from the solid phase unit storage module; when the sample moves to the position II, sample adding is carried out; and when the cleaning transfer module moves to the position III, the new sample adding position of the new sample adding transfer module is aligned with the new sample adding position of the cleaning transfer module in parallel.

The cleaning and transferring module moves in a linear reciprocating mode, when the cleaning and transferring module moves to a first position, a sample position to be tested of the cleaning and transferring module is aligned with a sample position to be tested of the incubation module in parallel, and a group of solid phase units are transferred from the incubation module to the cleaning and transferring module through the second gripper along the linear motion; when the solid phase unit of the cleaning and transferring module finishes adding the washing liquid and moves to the second position after cleaning, the sample position to be tested of the cleaning and transferring module is aligned with the sample position to be tested of the sample transferring module in parallel, and the solid phase unit on the sample position to be tested of the cleaning and transferring module is transferred to the sample position to be tested of the sample transferring module along the linear motion, and the solid phase unit to be tested is transferred to the measuring module for detection by the sample transferring module; when the cleaning and transferring module moves to a third position, the new sample adding position of the cleaning and transferring module is aligned with the new sample adding position of the new sample adding and transferring module positioned at the position III in parallel, and the third gripper transfers the solid phase unit on the new sample adding position of the new sample adding and transferring module to the new sample adding position of the cleaning and transferring module along the linear motion; when the cleaning and transferring module moves to the fourth position, the new sample adding position of the cleaning and transferring module is aligned with the new sample adding position of the incubation module in parallel, and the second gripper transfers the solid phase unit on the cleaning and transferring module to the incubation module for incubation along the linear motion.

In some preferred forms, the loading is automated by a sample charger.

In some preferred forms, the washing module includes a washing transfer module placement station to which the washing transfer module transfers solid phase units for placement and washing operations. The cleaning and transferring module directly moves into the cleaning module to carry out the arrangement and cleaning operation, so that the solid phase units in each reaction stage can be conveniently and directly cleaned simultaneously, and the operation is convenient and simple.

In some preferred embodiments, a plurality of analyte detection ligands are disposed within the solid phase unit, allowing for simultaneous detection of multiple analytes.

The invention has the beneficial effects that:

1. the qualitative and quantitative determination method of the analyte can realize the automation of the whole process from sample adding to detection of the analyzed sample, and has simple and convenient operation and high efficiency.

2. The invention adopts the solid phase unit to measure the analytes, the solid phase unit can be used singly, and can detect a plurality of analytes, and the operation is simpler and more, thereby being beneficial to keeping the consistency of the processing conditions.

3. By adopting the qualitative and quantitative determination method of the analyte, the single sample can be sequentially detected, the technical problems of inconsistent sample processing conditions and inaccurate determination results caused by simultaneous processing of a large number of samples are solved, and the sample determination accuracy is improved.

4. The qualitative and quantitative determination method of the analyte is based on the fact that incubation reaction and cleaning operation are needed in each step in the multi-step reaction, so that the simultaneous incubation and simultaneous cleaning of solid phase units in different reaction stages are realized, and the detection efficiency is greatly improved.

5. The qualitative and quantitative determination method of the analyte can realize the incubation and cleaning operation without interval among all groups of solid phase units, reduce the flow time and improve the detection efficiency.

Drawings

FIG. 1 is a top view of an assay system; a 'V' -shaped structure

FIG. 2 is a perspective view of an assay system;

FIG. 3 is a perspective view of a long plate incubation module;

fig. 4.1-4.3 are front, bottom and perspective views, respectively, of an embodiment of a disc-shaped incubation module;

FIGS. 5.1-5.2 are bottom and perspective views, respectively, of another embodiment of a disc-shaped incubation module;

FIG. 6 is a simplified top view of an assay system;

FIG. 7 is a top view of an embodiment of a cleaning transfer module in a first position;

FIG. 8 is a top view of an embodiment of a cleaning transfer module in a second position;

FIG. 9 is a top view of an embodiment of a cleaning transfer module in a third position;

FIG. 10 is a top view of an embodiment of a cleaning transfer module in a fourth position;

FIG. 11 is a simplified perspective view of an assay system;

FIG. 12 is a top view of another embodiment cleaning transfer module in a first position;

FIG. 13 is a top view of another embodiment cleaning transfer module in a second position;

FIG. 14 is a top view of another embodiment cleaning transfer module in a third position.

Detailed Description

For easy understanding, first, some technical terms related to the present invention will be further described:

analyte: substances capable of specifically binding to specific ligands include, but are not limited to, various types of antigens, antibodies, polypeptides, proteins, nucleic acids, and the like.

Solid phase unit: the solid phase unit is a container capable of bearing liquid, and a ligand lattice which is specifically combined with an analyte (sample) is immobilized on the inner side surface of the solid phase unit.

Three-step method, two-step method:

the enzyme-labeled substrate solution in the above table is only one of the exemplified measurement methods, and is not limited thereto.

In order that the objects, features and advantages of the invention will be readily understood, a more particular description of the invention will be rendered by reference to specific embodiments thereof which are illustrated in the appended drawings.

A qualitative and quantitative determination method for analytes comprises automatic sample loading, incubation, transportation, cleaning and detection.

Automatic sample feeding: the solid phase unit 110 is transferred from the solid phase unit storage module 120 to the new loading position 1811 of the new loading transfer module 181 by the first gripper, the new loading transfer module 181 is moved to the loading position, and the sample stored in the sample storage module 130 is sucked up by the sample filler 191 (shown in fig. 11) and then injected into the solid phase unit 110 on the new loading position 1811 of the new loading transfer module 181.

Incubation: incubation is performed on an incubation module 150, the incubation module 150 comprising an incubation tray 151, the incubation tray 151 having a solid phase unit receiving space disposed thereon. When the solid phase unit is inserted into the solid phase unit accommodating position, the temperature of the liquid inside the solid phase unit is regulated by the temperature inside the solid phase unit accommodating position, and the incubation is realized after a period of constant temperature treatment. Upon incubation, the ligand-specific reaction in the solid phase unit will reach equilibrium. By equilibrium is meant that the reaction between ligands does not change significantly in amount as the incubation time continues to extend.

Generally, in a certain temperature range, the reaction of ligand specific binding is characterized by a higher reaction temperature and a shorter time required to reach equilibrium, just as the reaction speed and temperature are related in chemical reactions. By applying the principle, the temperature in the holes of the incubation module can be regulated to increase the reaction speed and shorten the reaction time.

In some embodiments, the incubation module 150 comprises a long plate-shaped incubation plate 151, and the long plate-shaped incubation plate 151 comprises N2 groups of solid phase unit accommodating positions 1510 arranged in parallel in the length direction of the long plate-shaped incubation plate 151, wherein N2 = T/T, wherein T is the interval time from adding a solid phase unit into the solid phase unit accommodating position of the incubation module to adding the solid phase unit accommodating position of the incubation module again, and T is the interval time from moving a group of solid phase units into the solid phase unit accommodating position on the incubation module to moving a next group of solid phase units into the solid phase unit accommodating position. Each group of solid phase unit accommodation sites comprises N1 solid phase unit accommodation sites which are arranged in a straight line, wherein N1 = 2N or 3N, N is preferably 1 or 2 or 3, N new sample addition sites and N sample to be detected are respectively arranged at two ends. Taking t=10 min, the interval from the time when one group of solid phase units on the incubation module moves into the solid phase unit accommodating position of the incubation module to the time when the next group of solid phase units moves into the solid phase unit accommodating position of the incubation module is t=0.5 min, n1=3n, n=1 as an example, as shown in fig. 1-3, the long plate-shaped incubation plate 151 contains N2=20 groups of solid phase unit accommodating positions 1510, each group of solid phase unit accommodating positions contains 3 solid phase unit accommodating positions in a linear arrangement, the left side of each group of solid phase unit accommodating positions is a new sample adding position 1511 for accommodating solid phase units in the first reaction in the three-step method, the middle accommodating position is used for accommodating solid phase units in the second reaction in the three-step method, and the right side is a sample to be detected position 1512 for accommodating solid phase units in the last reaction in the three-step method. Therefore, the first 20 solid phase unit accommodation sites of the long plate-shaped incubation plate are all new sample addition sites 1511, and the third 20 solid phase unit accommodation sites are all sample to be measured.

The solid phase units after sample addition are added to the incubation plate 151 of the incubation module 150 at intervals of 0.5min, the 1 st solid phase unit is placed on the 1 st new sample addition position in the first row of the incubation plate, the 2 nd solid phase unit is placed on the 2 nd new sample addition position in the first row of the incubation plate, and until the 20 th solid phase unit is placed on the 20 th new sample addition position in the first row of the incubation plate. After the 20 th solid phase unit is placed, the 1 st solid phase unit just completes the first incubation reaction, the 1 st solid phase unit is moved out to be washed and added with a second step reaction reagent, then moved into the second row 1 st solid phase unit containing position of the incubation module, meanwhile, the 21 st solid phase unit is placed on the first row 1 st new sample adding position, the process takes 0.5min, the 2 nd solid phase unit completes incubation at the moment, is washed and added with the second step reaction reagent, then moved into the second row 2 nd solid phase unit containing position of the incubation module, meanwhile, the 22 nd solid phase unit is placed on the first row 2 nd new sample adding position, and the like in sequence until the 40 th solid phase unit is placed on the 20 th new sample adding position of the first row of the incubation tray. After the 40 th solid phase unit is placed, the 1 st and 21 st solid phase units just complete the second incubation reaction and the first incubation reaction respectively, the 1 st and 21 st solid phase units are simultaneously moved out for cleaning and adding the third step reaction reagent and the second step reaction reagent respectively, and then are respectively moved into the 1 st and the 1 st solid phase unit containing positions of the third row and the second row of the incubation module, and meanwhile, the 41 st solid phase unit is placed on the 1 st new sample adding position of the first row, and the process takes 0.5min. And so on until the 60 th solid phase unit is placed on the 20 th new sample adding position in the first column of the incubation plate. After the 60 th solid phase unit is placed, the 1 st, 21 st and 41 st solid phase units just complete the third, second and first incubation reactions respectively, the 1 st, 21 st and 41 st solid phase units are simultaneously moved out for cleaning, after the cleaning is completed, the 1 st solid phase unit is detected, the 21 st and 41 st solid phase units are respectively added with the reaction reagents of the third step and the second step, then are respectively moved into the holding positions of the 1 st and the 1 st solid phase units of the third row of the incubation module, and meanwhile, the 61 st solid phase unit is placed on the 1 st new sample adding position of the first row, the process takes 0.5min, and then the 2 nd, 22 nd and 42 nd solid phase units are subjected to related operations and are sequentially circulated. Therefore, single sample sequential detection is realized, and a group of solid phase units in each reaction stage of the three-step method are simultaneously incubated and cleaned, so that the detection efficiency is greatly improved.

In some embodiments, taking t=10 min, taking an example of the period from when one set of solid phase units on the incubation module moves into the incubation module solid phase unit accommodation space to when the next set of solid phase units moves into the incubation module solid phase unit accommodation space, t=0.5 min, n1=2n, n=1. The long plate-shaped incubation plate 151 contains N2 = 20 sets of solid phase unit accommodation sites 1510 arranged in parallel, each set of solid phase unit accommodation sites contains 2 solid phase unit accommodation sites arranged in a straight line, a new sample addition site 1511 is arranged on the left side of each set of solid phase unit accommodation sites for accommodating solid phase units in a first-step reaction in a two-step process, and a sample to be detected site 1512 is arranged on the right side for accommodating solid phase units in a second-step reaction in the two-step process. Therefore, the first 20 solid phase unit accommodation sites of the long plate-shaped incubation plate are all new sample addition sites 1511, and the second 20 solid phase unit accommodation sites are all sample to be measured.

The solid phase units after sample addition are added to the incubation plate 151 of the incubation module 150 at intervals of 0.5min, the 1 st solid phase unit is placed on the 1 st new sample addition position in the first row of the incubation plate, the 2 nd solid phase unit is placed on the 2 nd new sample addition position in the first row of the incubation plate, and until the 20 th solid phase unit is placed on the 20 th new sample addition position in the first row of the incubation plate. After the 20 th solid phase unit is placed, the 1 st solid phase unit just completes the first incubation reaction, the 1 st solid phase unit is moved out to be washed and added with a second reaction reagent, then moved into the second row of 1 st sample to be tested of the incubation module, meanwhile, the 21 st solid phase unit is placed on the 1 st new sample adding position of the first row, the process takes 0.5min, the 2 nd solid phase unit completes incubation at the moment, the washing and the second reaction reagent are carried out, then moved into the second row of 2 nd sample to be tested of the incubation module, meanwhile, the 22 nd solid phase unit is placed on the 2 nd new sample adding position of the first row, and the like are sequentially carried out until the 40 th solid phase unit is placed on the 20 th new sample adding position of the first row of the incubation tray. After the 40 th solid phase unit is placed, the 1 st and 21 st solid phase units just complete the second incubation reaction and the first incubation reaction respectively, the 1 st and 21 st solid phase units are simultaneously moved out for cleaning, after the cleaning is completed, the 1 st solid phase unit is detected, the 21 st solid phase unit is added with a second step reaction reagent and then moved into the second row of the incubation module, the 1 st sample to be detected is placed on the 1 st new sample adding position of the first row, the process takes 0.5min, then the 2 nd and 22 nd solid phase units are subjected to related operations, and the process is circulated in sequence. Therefore, single sample sequential detection is realized, and a group of solid phase units at each reaction stage in the two-step method are simultaneously incubated and cleaned, so that the detection efficiency is greatly improved.

In some embodiments, as shown in fig. 4.1-5.2, the incubation module 150 'comprises a disc-shaped incubation plate 151' on which at least one set of solid phase unit accommodation sites are arranged in a circumferential direction, each set of solid phase unit accommodation sites is arranged in a straight line, a line connecting a midpoint of the line with a center of the circle is perpendicular to a line connecting each set of solid phase unit accommodation sites, or an extension line of the line connecting each set of solid phase unit accommodation sites passes through the center of the circle. The incubation of the solid phase units on the disc-shaped incubation plate 151' is similar to that on the long plate-shaped incubation plate, after a group of solid phase units are washed and moved into the accommodation position of the solid phase units of the incubation module, the rotating device 152' at the bottom of the disc-shaped incubation plate 151' drives the disc-shaped incubation plate 151' to rotate, and the cover body 153' is not moved, so that the next group of solid phase units are exposed out of the opening 154' of the cover body 153' for the next operation.

Transport of: with automatic transfer, the automatic transfer is performed by a transfer module, as shown in fig. 6, where the transfer module includes a new sample adding transfer module 181, a cleaning transfer module 182, a to-be-tested sample transfer module 183, a first gripper 184, a second gripper 185, and a third gripper 186, and the new sample adding transfer module 181 includes N3 new sample adding positions 1811 of solid phase units, where n3=n; the sample transfer module 183 comprises N4 solid phase unit sample sites 1831, where n4=n.

In some embodiments, the wash transfer module comprises a set of N5 in-line solid phase unit receptacles, where n5=n1+n, comprising N solid phase unit fresh sample addition stations and N solid phase unit test samples at both ends. Taking n=1 as an example, as shown in fig. 6, each group of metal incubation plates 151 in the incubation module 150 includes 3 solid phase unit accommodation sites, the leftmost new sample addition site 1511, and the rightmost sample to be tested site 1512; the cleaning and transferring module 182 includes 4 accommodation positions, namely a new sample loading position 1821 on the leftmost side and a sample to be tested position 1822 on the rightmost side; the fresh sample transfer module 181 comprises 1 fresh sample site 1811 and the sample transfer module 183 comprises 1 sample site 1831. The cleaning transfer module 182, the new sample loading transfer module 181 and the sample transfer module 183 to be tested all reciprocate along a straight line, and the first gripper 184 has 1 solid phase unit gripping position and can reciprocate along the straight line. The second gripper 185 has 3 solid phase unit gripping sites and can reciprocate linearly along the length of the incubation plate 151. The third gripper 186 has 1 solid phase unit gripping position and reciprocates in a straight line.

The cleaning and transferring module includes 4 positions along the linear motion, and is located at the first position, as shown in fig. 7, the sample position 1822 to be tested of the cleaning and transferring module 182 is aligned with the sample position 1512 to be tested of the incubation module 150 in parallel, at this time, the second gripper 185 grabs 3 solid phase units in different reaction phases on the incubation module 150, and moves along the linear motion to the position right above the cleaning and transferring module 182, so that the solid phase units on the incubation module 150 are aligned one by one and transferred to the accommodating position of the cleaning and transferring module 182. The wash transfer module 182 then transfers the solid phase units to the wash module 160, completing the wash and rinse, during which the first gripper 184 grips the solid phase units 110 in the solid phase storage module 120 and transfers them in a straight line to the fresh sample application station 1811 of the fresh sample application transfer module 181 at station I, and then the fresh sample application transfer module 181 moves to station II for application of samples. The wash transfer module 182 is then transferred to a second position, as shown in fig. 8, while the sample transfer module 183 is moved from within the measurement module 170 to outside the measurement module, the wash transfer module 182 sample position 1822 (below the third gripper 186, not shown in fig. 8) is aligned parallel to the sample position 1831 of the sample transfer module 183, the third gripper 186 grabs the solid phase unit on the sample position 1822 of the wash transfer module 182 and transfers the solid phase unit to the sample position 1831 of the sample transfer module 183 in a straight line, and the sample transfer module 183 transfers the solid phase unit to the measurement module 170 in a straight line for measurement. When the wash transfer module 182 is moved to the third position, as shown in fig. 9, the solid phase unit after completing the loading is transferred to the position iii by the new loading transfer module 181, and at this time, the new loading position 1821 of the wash transfer module 182 is aligned with the new loading position 1811 of the new loading transfer module 181 located at the position iii (the new loading position 1821 of the wash transfer module 182 and the new loading position 1811 of the new loading transfer module 181 are located under the third gripper 186, which is not shown in fig. 9), and the third gripper 186 grips the solid phase unit from the new loading position 1811 of the new loading transfer module 181 and transfers to the new loading position 1821 of the wash transfer module 182. The wash transfer module 182 is then moved to a fourth position, as shown in fig. 10, where the wash transfer module 182 new loading position 1821 is aligned parallel to the incubation module 150 new loading position 1511 (the wash transfer module 182 new loading position 1821 is located below the second gripper 185, not shown in fig. 10), and the second gripper 185 grabs and transfers the solid phase units on the wash transfer module 182 onto the incubation module 150 in one-to-one alignment, and the solid phase units on the post-transfer wash transfer module 182 new loading position 1821 are located on the incubation module 150 new loading position 1511. Then, the second gripper grips the next set of solid phase units for the above operation and circulates in sequence.

In some embodiments, the washing transfer module 182 includes a set of N5 solid phase unit receptacles aligned in a line, where n5=n1, i.e., the number of solid phase unit receptacles of the washing transfer module 182 is the same as the number of solid phase unit receptacles of the incubation module 150, and the solid phase unit receptacles of the washing transfer module include N solid phase unit test sample receptacles 1512 at one end. Taking n=1 as an example, as shown in fig. 12-14, each group of incubation modules 150 includes 3 solid phase unit accommodation sites, the leftmost new sample addition site 1511, and the rightmost sample measurement site 1512; the cleaning and transferring module 182 includes 3 accommodation sites, and the rightmost site is a sample site 1822 to be tested; the fresh sample transfer module 181 comprises 1 fresh sample site 1811 and the sample transfer module 183 comprises 1 sample site 1831. The cleaning transfer module 182, the new sample adding transfer module 181 and the to-be-tested sample transfer module 183 all reciprocate along a straight line, and the first gripper 184 is located right above the solid phase unit in the solid phase unit storage module 120, has 1 solid phase unit grabbing position, and can reciprocate along a straight line. The second gripper 185 is located right above the long plate-shaped incubation plate 151 in the incubation module 150, has 3 solid phase unit gripping positions, and can linearly reciprocate along the length direction of the incubation plate 151. The third gripper 186 has 1 solid phase unit gripping position and reciprocates in a straight line.

The cleaning and transferring module 182 includes 3 positions along the linear motion, and when the cleaning and transferring module 182 is located at the first position, as shown in fig. 12, the sample position 1822 to be tested of the cleaning and transferring module 182 is aligned with the sample position 1512 to be tested of the incubation module 150 in parallel, at this time, the second gripper 185 grabs 3 solid phase units in different reaction phases on the incubation module 150, and moves along the linear motion to right above the cleaning and transferring module 182, so that the solid phase units are aligned and transferred onto the cleaning and transferring module 182, and the solid phase units on the sample position 1512 to be tested of the incubation module 150 after transfer are located on the sample position 1822 to be tested of the cleaning and transferring module 182. The wash transfer module 182 then transfers the solid phase units to the wash module 160, and the washing and washing is completed, during which the first gripper 184 grabs the solid phase unit 110 in the solid phase storage module 120 and transfers it along a line to the fresh sample transfer module 181 at position i at the fresh sample transfer position 1811, and then the fresh sample transfer module 181 moves to position ii where the sample is transferred, at which time the fresh sample transfer module 181 and the wash transfer module 182 are on the same line, and the fresh sample transfer module 181 at the fresh sample transfer position 1811 is offset from the fresh sample transfer position 1511 of the incubation module 150 in parallel. The wash transfer module 182 is then moved to a second position (as shown in fig. 13) while the sample transfer module 183 is moved from within the measurement module 170 to outside the measurement module 170. The wash transfer module 182 sample position 1822 (below the third hand 186, not shown in fig. 13) is aligned parallel to the sample position 1831 of the sample transfer module 183. The third hand 186 grabs the solid phase unit on the sample position 1822 of the wash transfer module 182 and linearly transfers to the sample position 1831 of the sample transfer module 183. The sample transfer module 183 transfers the solid phase unit to the measurement module 170 for measurement. When the wash transfer module 182 moves to the third position, as shown in fig. 14, the completed new solid phase unit is transferred to the position iii by the new transfer module 181, at this time, the new transfer module 181 and the wash transfer module 182 are located in the same straight line, and the new sample adding position 1811 of the new transfer module 181 is aligned with the new sample adding position 1511 of the incubation module 150 (the new sample adding position 1811 of the new transfer module 181 is located below the second grip 185, not shown in fig. 14), the new sample adding position 1811 of the new transfer module 181 is adjacent to the wash transfer module 182, the second grip 185 grabs and transfers the new sample adding position 1811 of the new sample adding module 181 and the solid phase unit on the wash transfer module 182 to the incubation module 150, and the solid phase unit on the new sample adding position 1811 of the new transfer module 181 after transfer is located on the new sample adding position 1511 of the incubation module 150. Then, the second gripper grips the next set of solid phase units for the above operation and circulates in sequence.

Cleaning: cleaning is performed on a cleaning module 160, the cleaning module 160 including a cleaning transfer module placement location. The cleaning transfer module transfers a group of solid phase units to move to a preset position, a cleaning liquid filler 193 (shown in fig. 11) fills the cleaning liquid into the solid phase units, the cleaning transfer module transfers the solid phase units to a position in the cleaning module 160 after the cleaning is completed, the cleaning liquid is centrifugally dried, and after the cleaning is completed once, the cleaning liquid filling and the centrifugal cleaning are repeatedly performed, wherein the cleaning times are not more than 3 times.

Measurement: the measurement is carried out on the measurement module, the solid phase unit is transported into the measurement module through the to-be-measured sample transporting module, substrate liquid is injected, and collection of signals at the bottom of the solid phase unit is started when the turntable rotates to the photographing objective lens opening. The collection will be completed within a certain time but not more than the time the turntable is rotated again.

Adding reagent: in some embodiments, as shown in fig. 11, reagents are injected into the solid phase cell by reagent injector 192, which for a three-step process includes a first reagent injector 1921 for injecting a second step reagent and a second reagent injector 1922 for injecting a third step reagent; the automatic filling of the reaction reagent can be realized through the reagent liquid charger.

Although the present invention is disclosed above, the present invention is not limited thereto. Various changes and modifications may be made by one skilled in the art without departing from the spirit and scope of the invention, and the scope of the invention should be assessed accordingly to that of the appended claims.

Claims

1. A qualitative and quantitative determination method for analytes, which is characterized by comprising automatic sample adding, incubation, cleaning and detection, wherein the incubation and the cleaning are respectively carried out on an incubation module and a cleaning module, a group of solid phase units in different reaction phases start and end incubation on the incubation module and start and end cleaning on the cleaning module simultaneously; the incubation module comprises solid phase unit accommodation positions which are formed by N2 and are arranged in a straight line, each group of solid phase unit accommodation positions comprises a new sample adding position and a sample to be tested at two ends of a branch line, and a group of solid phase units in each reaction stage are placed in the corresponding accommodation positions for incubation reaction; and the N2 = T/T, wherein T is the interval time from adding the solid phase unit into the solid phase unit accommodating position of the incubation module to adding the solid phase unit accommodating position of the incubation module again, and T is the interval time from moving one group of solid phase units into the solid phase unit accommodating position of the incubation module to moving the next group of solid phase units into the solid phase unit accommodating position of the incubation module.

2. The method of claim 1, wherein each set of solid phase unit receptacles of the incubation module comprises N1 solid phase unit receptacles, wherein n1=2n or 3N, N new sample receptacles and N test receptacles are arranged at two ends.

3. The method of claim 2, wherein after a set of solid phase units is washed, the solid phase units on the sample position to be detected of the original incubation module are detected, the rest solid phase units are transferred to the incubation module and are shifted from the new sample adding position to the direction of the sample position to be detected on the incubation module, and meanwhile, the new sample adding position is supplemented with the new solid phase units after sample adding, and the incubation is performed again.

4. A method according to claim 3, wherein after the last set of solid phase units has been washed and incubation is resumed, the next set of solid phase units is washed, incubated again until the last set, and cycled through.

5. The method of claim 4, wherein the incubation module comprises an elongated plate-shaped incubation plate on which each set of solid phase unit receptacles are arranged in parallel along the length.

6. The method of claim 4, wherein the incubation module comprises a disc-shaped incubation disc on which each set of solid phase unit receptacles are arranged in a circumferential direction.

7. The method of claim 6, wherein the line between the midpoint of the line between each set of solid phase cell accommodation sites and the center of the circle is perpendicular to the line between each set of solid phase cell accommodation sites or the extension line of the line between each set of solid phase cell accommodation sites passes through the center of the circle.

8. The method of claim 6, wherein the incubation plate is provided with a rotating means at the bottom and a cover having openings exposing a set of solid phase units.

9. The method of claim 8, wherein after the previous set of solid phase units is washed and incubation is resumed, the rotating device drives the incubation plate to rotate while the cover is stationary, and the next set of solid phase units is exposed through the cover opening for washing and incubation steps.

10. The method of any one of claims 1-9, further comprising an automated transfer step, the automated transfer being accomplished by a transfer module, the transfer module comprising a wash transfer module, a fresh transfer module, a test sample transfer module, a first gripper, a second gripper, a third gripper; the new sample loading transfer module comprises N3 new sample loading positions, and the sample to be tested transfer module comprises N4 sample to be tested positions, wherein N3=N4=n.

11. The method of claim 10, wherein the wash transfer module comprises a set of N5 in-line solid phase unit receptacles, wherein n5=n1, comprising N solid phase unit test sites at one end.

12. The method of claim 11, wherein the fresh sample transfer module is positioned in a straight line with the wash transfer module, the fresh sample transfer module reciprocates in a straight line, and the first gripper transfers the solid phase unit from the solid phase unit storage module to a fresh sample application position of the fresh sample transfer module when the fresh sample transfer module moves to position I; when the sample moves to the position II, sample adding is carried out; and when the sample is moved to the position III, the new sample adding position of the new sample adding transfer module is aligned with the new sample adding position of the incubation module in parallel.

13. The method of claim 12, wherein the wash transfer module reciprocates in a straight line and the wash transfer module is moved to a first position with the sample position of the wash transfer module aligned parallel to the sample position of the incubation module and the second gripper transfers a set of solid phase units from the incubation module to the wash transfer module; when the solid phase unit of the cleaning and transferring module finishes adding the washing liquid and moves to the second position after cleaning, the sample position to be tested of the cleaning and transferring module is aligned with the sample position to be tested of the sample transferring module in parallel, and the solid phase unit on the sample position to be tested of the cleaning and transferring module is transferred to the sample position to be tested of the sample transferring module by the third gripper, and the solid phase unit to be tested is transferred to the measuring module by the sample transferring module to be tested for detection; when the cleaning and transferring module moves to the third position, the cleaning and transferring module is closely adjacent to the new sample adding and transferring module located at the position III, and the second gripper transfers the new sample adding and transferring module and the solid phase unit on the cleaning and transferring module to the incubation module.

14. The method of claim 10, wherein the wash transfer module comprises a set of N5 in-line solid phase unit receptacles, wherein n5=n1+n, comprising N solid phase unit fresh sample sites and N solid phase unit test sample sites at both ends.

15. The method of claim 14, wherein the fresh sample transfer module reciprocates in a straight line and, when moved to position I, the first gripper transfers the solid phase unit from the solid phase unit storage module to the fresh sample transfer module at a fresh sample application position; when the sample moves to the position II, sample adding is carried out; and when the cleaning transfer module moves to the position III, the new sample adding position of the new sample adding transfer module is aligned with the new sample adding position of the cleaning transfer module in parallel.

16. The method of claim 15, wherein the wash transfer module reciprocates in a straight line and the wash transfer module is moved to a first position with the sample position of the wash transfer module aligned parallel to the sample position of the incubation module and the second gripper transfers a set of solid phase units from the incubation module to the wash transfer module; when the solid phase unit of the cleaning and transferring module finishes adding the washing liquid and moves to the second position after cleaning, the sample position to be tested of the cleaning and transferring module is aligned with the sample position to be tested of the sample transferring module in parallel, and the solid phase unit on the sample position to be tested of the cleaning and transferring module is transferred to the sample position to be tested of the sample transferring module by the third gripper, and the solid phase unit to be tested is transferred to the measuring module by the sample transferring module to be tested for detection; when the cleaning and transferring module moves to a third position, the new sample adding position of the cleaning and transferring module is aligned with the new sample adding position of the new sample adding and transferring module positioned at the position III in parallel, and the third gripper transfers the solid phase unit on the new sample adding position of the new sample adding and transferring module to the new sample adding position of the cleaning and transferring module; when the cleaning and transferring module moves to the fourth position, the new sample adding position of the cleaning and transferring module is aligned with the new sample adding position of the incubation module in parallel, and the second gripper transfers the solid phase unit on the cleaning and transferring module to the incubation module for incubation.

17. The method of claim 1, wherein the loading is automated by a sample applicator.

18. The method of claim 10, wherein the washing module comprises a washing transfer module positioning station to which the washing transfer module transfers the solid phase units for positioning and washing operations.

19. The method of claim 1, wherein a plurality of analyte detection ligands are disposed within the solid phase unit.