CN116068186A

CN116068186A - Method, system and application for identifying HOOK effect sample in immunoassay

Info

Publication number: CN116068186A
Application number: CN202111285038.4A
Authority: CN
Inventors: 韩双亭; 张黎明; 李临
Original assignee: Kemei Boyang Diagnostic Technology Shanghai Co ltd
Current assignee: Kemei Boyang Diagnostic Technology Shanghai Co ltd
Priority date: 2021-11-01
Filing date: 2021-11-01
Publication date: 2023-05-05

Abstract

The present invention relates to a method and system for identifying a sample of the HOOK effect in an immunoassay. The method comprises the following steps: s1, carrying out two parallel immune response detection on a sample to be detected containing target molecules to be detected, recording detection results of the two parallel immune responses, and respectively calculating a first detection value and a second detection value; s2, calculating a ratio of the first measured value to the second measured value; s3, acquiring a critical point c of a standard substance, wherein detection results of two parallel immune reactions of the standard substance corresponding to the critical point c are respectively calculated as a measured value a and a measured value a ', and the measured value a is larger than the measured value a'; s4, comparing the ratio of the first measured value to the second measured value with the critical point c, and further identifying whether the sample to be tested is a HOOK effect sample. The method can solve the problem that the traditional detection reagent is difficult to distinguish the HOOK effect sample, and is not limited by the detection range.

Description

Method, system and application for identifying HOOK effect sample in immunoassay

Technical Field

The invention belongs to the technical field of immunodetection, and particularly relates to a method, a system and application for identifying a HOOK effect sample in immunoassay.

Background

Immunological detection is based on the principle of antigen-antibody specific reaction, and is often used for detecting trace amounts of bioactive substances such as proteins and hormones, because it can display the analyte or amplify the signal by using isotopes, enzymes, chemiluminescent substances, and the like.

The photoexcitation chemiluminescence method is one of the common methods of chemiluminescence analysis technology, can be used for researching the interaction between biomolecules, and is mainly used for detecting diseases clinically. The technology integrates the research of the related fields of polymer particle technology, organic synthesis, protein chemistry, clinical detection and the like. The technical principle of the photoexcitation chemiluminescence analysis technology is that a sensitizer can excite oxygen molecules in the surrounding environment into singlet oxygen molecules under the irradiation of laser, and the singlet oxygen molecules can react with a luminous composition with the distance of about 200nm to generate a light signal with a certain wavelength; when the sample contains the antigen or antibody to be detected, the immune reaction of the antigen and the antibody can enable the donor particles containing the sensitizer to be combined with the acceptor particles containing the luminous composition, so that a light signal with a specific wavelength is generated, and the content of the antigen or antibody to be detected can be detected by detecting the light signal.

In the antigen-antibody dose response curve, when the amount of antibody is fixed, the response signal shows a phenomenon of rising and then falling with the increase of the amount of antigen. The region in which the response signal rises with increasing antigen dose is referred to as the "front band" region, the region in which the response signal falls with increasing antigen dose is referred to as the "rear band" region, and the region where the front band and rear band are joined is referred to as the "equivalent band".

In the immune reaction, the reactivity of the antigen and the antibody shows a phenomenon of rising and then falling as the ratio of the antigen to the antibody rises, and is called a "HOOK effect" or a "HOOK effect". Clinically, the hook effect can lead to high value samples producing false negative results, i.e. "false negatives".

The current immunoassay method generally uses the front band region of the dose-response curve to calculate the concentration of the analyte according to the linear relationship between the analyte content and the response signal. This detection method has a number of drawbacks, such as:

because the traditional immunodetection reagent lacks a means for identifying the HOOK effect, a clinician is often required to identify whether the HOOK effect exists in the sample by adopting a method for diluting a serum sample in combination with the clinical manifestation of a patient, and the traditional immunodetection reagent is complex in operation, long in time consumption and easy to cause missed detection.

Disclosure of Invention

In view of the shortcomings of the prior art, it is an object of the present invention to provide a method, system and application for identifying a sample of the HOOK effect in an immunoassay. By using the method and the system, whether the sample to be detected is the HOOK effect sample can be simply, conveniently, quickly and accurately identified.

In order to achieve the above object and other related objects, the present invention adopts the following technical scheme:

in a first aspect the present invention provides a method of identifying a sample of the HOOK effect in an immunoassay comprising the steps of:

s1, carrying out two parallel immune response detection on a sample to be detected containing target molecules to be detected, recording detection results of the two parallel immune responses, and respectively calculating a first detection value and a second detection value;

s2, calculating a ratio of the first measured value to the second measured value;

s3, acquiring a critical point c of a standard substance, wherein detection results of two parallel immune reactions of the standard substance corresponding to the critical point c are respectively calculated as a measured value a and a measured value a ', and the measured value a is larger than the measured value a';

s4, comparing the ratio of the first measured value to the second measured value with the critical point c, and further identifying whether the sample to be tested is a HOOK effect sample.

In some embodiments of the present invention, when the ratio of the first measured value to the second measured value is smaller than the critical point c, the sample to be tested is a non-HOOK effect sample;

when the ratio of the first measured value to the second measured value is greater than the critical point c and the first measured value is greater than the measured value a, the sample to be measured is a sample with non-HOOK effect.

In other embodiments of the present invention, when the ratio of the first measured value to the second measured value is greater than the critical point c and the first measured value is smaller than the measured value a, the sample to be tested is a HOOK effect sample.

In some embodiments of the invention, the ratio of the content of target molecule to be detected/the content of specific capture molecule is different in the two parallel immunoreactions; wherein the specific capture molecule is capable of specifically binding to the target molecule to be detected.

In some embodiments of the invention, in the two parallel immunoreaction assays, the assay result of an immunoreaction with a greater ratio of the content of the specific capture molecule to the content of the target molecule to be assayed is a first assay or assay value a, and the other is a second assay or assay value a'.

In some embodiments of the present invention, in step S2, the method for obtaining the critical point c includes the following steps:

a1, detecting a series of standard substances with known target molecule content and different concentrations, wherein each standard substance is subjected to two parallel immunoreaction detection, and the detection results of the two parallel immunoreactions are recorded and respectively calculated as a measured value a and a measured value a';

a2, a reaction curve A of the measured value a and the concentration of the standard substance is made and stored;

a3, a reaction curve B of the measured value a' and the concentration of the standard substance is made and stored;

and A4, taking a point from the overlapping part of the standard substance concentration corresponding to the front zone area of the reaction curve A and the rear zone area of the reaction curve B, and recording the ratio of the measured value a/the measured value a' corresponding to the point as a critical point c and storing the critical point c.

In other embodiments of the invention, the two parallel immunoreactions are detected with a first measurement and a second measurement for an immunoreaction with a greater ratio of the amount of specific capture molecule to the amount of target molecule to be detected.

In some embodiments of the invention, the target molecule to be tested is selected from an antigen or an antibody; and/or

The specific capture molecules include a first capture molecule bound to a solid phase material and a second capture molecule labeled with a label.

In other embodiments of the invention, the specific capture molecule is present in an amount that is the amount of the first capture molecule, the amount of the second capture molecule, or the sum of the amounts of the first and second capture molecules; preferably the content of the first capture molecules.

In a second aspect the invention provides a system for carrying out the method according to the first aspect of the invention, comprising:

the immune reaction device is provided with a reaction cup, and the reaction cup is used for loading a sample to be tested and a detection reagent to perform immune reaction;

the immunoassay device is provided with a data acquisition device;

the controller is used for moving two reaction cups filled with the same sample to be detected and corresponding detection reagents in the immune reaction device to the immune determination device, the data acquisition device acquires data of two parallel immune reaction detections of the same sample to be detected and evaluates the acquired data to obtain detection results of the two parallel immune reactions, and the detection results are respectively calculated to be a first measurement value and a second measurement value;

and the processor is used for calculating the ratio of the first measured value to the second measured value and identifying whether the sample to be tested is the HOOK effect sample or not.

In some embodiments of the invention, the two reaction cups for carrying the same sample to be tested and the corresponding detection reagents are placed at any given position on the immunoreaction apparatus; preferably, the two reaction cups for carrying the same sample to be tested and the corresponding detection reagents are positioned adjacent to each other on the immunoreaction device.

In other embodiments of the invention, the immunoassay device performs data acquisition on two parallel immunoreaction assays of the same sample to be tested sequentially; preferably, the immunoassay device performs data acquisition on two parallel immunoreaction detections of the same sample to be tested at the same time.

In some embodiments of the present invention, the processor stores a reaction curve a, a reaction curve B, and a critical point c, for identifying whether the sample to be tested is a HOOK effect sample.

In a third aspect, the invention provides the use of a method according to the first aspect or a system according to the second aspect of the invention in chemiluminescent immunoassay, enzyme linked immunoassay and immunonephelometric immunoassay; preferably in chemiluminescent immunoassay; further preferred is the use in photo-activated chemiluminescence immunoassay.

Compared with the prior art, the invention has the beneficial effects that:

the method and the system can simply, conveniently, quickly and accurately identify whether the sample to be detected is the HOOK effect sample, avoid missing detection caused by the HOOK effect, and are not limited by the detection range.

Drawings

Figure 1 is a dose response curve for antigen-antibody.

Fig. 2 is a schematic diagram of the method of the present invention.

FIG. 3 is a graph showing the reaction of standard substance concentration with reagent 1 signal and reagent 2 signal in example 2.

Fig. 4 is a schematic diagram of the system according to the present invention.

Detailed Description

In order that the invention may be readily understood, the invention will be described in detail. Before the present invention is described in detail, it is to be understood that this invention is not limited to particular embodiments described. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.

Where a range of values is provided, it is understood that each intervening value, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the invention, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

Unless defined otherwise, all terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention, the preferred methods and materials are now described.

According to the method for identifying the HOOK effect sample in the immunoassay, two parallel tests are carried out on each sample, the ratio of the content of the target molecule to be detected to the content of the specific capture molecule in the two tests is different, and finally two different signals are generated, namely a first measured value and a second measured value. As the content of the target molecule to be detected increases, the ratio of the first measured value to the second measured value continuously rises and shows a certain linear relationship. According to the principle, the method for identifying the HOOK effect sample in the immunoassay is as follows:

using the critical point c to identify whether the sample to be tested is a HOOK effect sample:

in the antigen-antibody dose response curve (fig. 1), when the amount of antibody is fixed, the response signal shows a phenomenon of ascending followed by descending with the increase of the amount of antigen. The region in which the reaction signal increases with increasing antigen amount is referred to as a front band region, the region in which the reaction signal decreases with increasing antigen amount is referred to as a rear band region, and the region in which the front band and the rear band are joined is referred to as an equivalent band.

As shown in fig. 2, the reaction curves a and B are reaction curves obtained from the first and second measured values and the standard substance concentration, respectively. In the portion where the front band region of the reaction curve a and the rear band region of the reaction curve B overlap in concentration (as in the portion in the dashed box in fig. 2), a point is taken in the above-mentioned portion where the concentrations overlap, and the ratio of the first measured value to the second measured value (as in a/b=15) corresponding to the point is taken as the critical point c.

When the ratio of the first measured value to the second measured value is smaller than the critical point c, the sample to be detected is a non-HOOK effect sample;

when the ratio of the first measured value to the second measured value is larger than the critical point c and the first measured value is larger than the measured value a, the sample to be detected is a sample with a non-HOOK effect;

when the ratio of the first measured value to the second measured value is greater than the critical point c and the first measured value is smaller than the measured value a, the sample to be measured is a HOOK effect sample.

In some embodiments of the present invention, the method for identifying a sample of the HOOK effect in an immunoassay provided by the present invention specifically comprises the steps of:

the critical point c is first obtained by a method comprising the steps of:

a1, detecting a series of standard substances with known target molecule content and different concentrations, wherein each standard substance is subjected to two parallel immunoreaction detection, and the detection results of the two parallel immunoreactions are recorded and respectively calculated as a measured value a and a measured value a'; the detection mode of the measured value a is the same as that of a first measured value of the sample to be detected, and the detection mode of the measured value a' is the same as that of a second measured value of the sample to be detected;

Then, whether the sample to be tested is a HOOK effect sample is identified by a method comprising the following steps:

s3, comparing the ratio of the first measured value to the second measured value with the critical point c, and when the ratio of the first measured value to the second measured value is smaller than the critical point c, the sample to be detected is a non-HOOK effect sample; when the ratio of the first measured value to the second measured value is larger than the critical point c and the first measured value is larger than the measured value a, the sample to be detected is a sample with a non-HOOK effect; when the ratio of the first measured value to the second measured value is greater than the critical point c and the first measured value is smaller than the measured value a, the sample to be measured is a HOOK effect sample.

According to some embodiments of the invention, the difference between the specific capture molecule content and the target molecule content can be achieved by referring to, but not limited to, the following ways:

in some embodiments of the invention, the two parallel immunoreactions are detected with a first measurement and a second measurement for immunoreactions having a greater ratio of the amount of specific capture molecule to the amount of target molecule to be detected.

The target molecule to be detected according to the present invention refers to any inorganic or organic molecule that can be detected immunologically, including any target biological substance. Examples representing the target molecules to be tested are cells, viruses, subcellular particles, proteins, lipoproteins, glycoproteins, peptides, polypeptides, nucleic acids, oligosaccharides, polysaccharides, lipopolysaccharides, cellular metabolites, haptens, hormones, pharmaceutical substances, alkaloids, steroids, vitamins, amino acids and sugars.

The specific capture molecule according to the present invention refers to a molecule capable of binding to another molecule (target molecule to be detected) due to the attractive interaction between the molecules. Examples of such specific capture molecules include, but are not limited to, proteins, nucleic acids, carbohydrates, lipids, and small organic molecules. By specific capture molecule is meant a capture molecule capable of recognizing and binding to a specific target molecule to be detected, but not any target molecule.

According to some embodiments of the invention, the target molecule to be tested is selected from an antigen or an antibody. According to some embodiments of the invention, the antigen is any substance having immunogenicity. Including but not limited to those listed in the examples of target molecules described above that are immunogenic. According to some embodiments of the invention, the specific capture molecule is selected from one member of a specific binding pair, such as an antibody, and the target molecule to be detected is the other member of the specific binding pair, such as an antigen to which it is paired. The term "antibody" is used herein in the broadest sense and specifically covers monoclonal antibodies, polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies) formed from at least two intact antibodies, and antibody fragments (e.g., fab regions, fc regions, single chain antibodies).

According to some embodiments of the invention, the specific capture molecules employed in the immunoreaction assays comprise a first capture molecule bound to a solid phase material and a second capture molecule labeled with a label. The first capture molecule and the second capture molecule may be the same or different, and may be the same or different, but are each capable of specifically binding to the target molecule to be detected.

According to some embodiments of the invention, the first capture molecule bound solid phase material is selected from the group consisting of particles, microparticles, beads, electrodes and multi-well plates. In some embodiments, the first capture molecule is a luminescent particle binding that contains a luminescent group that is capable of rapidly absorbing singlet oxygen and then emitting light at a wavelength (e.g., 500-615 nm).

The standard substance refers to a target molecule solution to be measured, the content of which is known or which can quantitatively determine and assign the target molecule to be measured.

The "content" of a substance according to the invention refers to the mass concentration of the substance in its respective solution.

A second aspect of the invention relates to a system for implementing the method according to the first aspect of the invention, comprising: the immune reaction device, the immune determination device, the controller and the processor can perform two parallel tests on each sample, and identify whether the sample to be tested is a HOOK effect sample according to the obtained two parallel test results.

In the invention, the immune reaction device is provided with a reaction cup, and the reaction cup is used for loading a sample to be tested and a detection reagent to carry out immune reaction. In the present invention, the form of the reaction cup is not particularly limited, and the reaction cup may be a single reaction cup or a plurality of reaction cups may be assembled together to form a strip (row) type or plate type reaction cup group. In the invention, two reaction cups for loading the same sample to be tested and corresponding detection reagents can be at any appointed position on the immunoreaction device. In some embodiments of the invention, two reaction cups for carrying the same sample to be tested and corresponding detection reagents are positioned adjacent to each other on the immunoreaction apparatus.

The immunoassay device is provided with a data acquisition device; the data collector can be a device for collecting optical signals, the optical signals can be optical signals emitted by the luminous composition after being irradiated by excitation light, optical signals emitted by the luminous composition after being excited by chemical energy, optical signals generated after electrochemical reaction on the surface of an electrode or optical signals transmitted through suspended particle media, and the like, so that the system can be applied to light excitation chemiluminescence immunoassay, fluorescence immunoassay, electrochemiluminescence immunoassay, enzyme-linked immunoassay, nephelometry immunoassay, and the like. The data collector can also evaluate the collected data.

In some embodiments of the invention, the system is used in a photoexcitation chemiluminescent immunoassay. At this time, the data collector is a photon counter, and further, the immunoassay device further comprises an optical exciter for emitting excitation light and irradiating the excitation light into the reaction cup, so as to excite a compound formed by immune reaction to emit an optical signal, and the photon counter receives the optical signal and records a reading.

According to the invention, the controller moves two reaction cups filled with the same sample to be detected and corresponding detection reagent in the immunoreaction device to the position of the immunoassay device according to a set instruction, and then the data acquisition device acquires data for two parallel immunoreaction detections of the same sample to be detected.

In some embodiments of the present invention, the immunoassay device performs data acquisition on two parallel immunoreaction assays of the same sample to be tested sequentially; in other embodiments of the invention, the immunoassay device simultaneously performs data collection for two parallel immunoreaction assays of the same test sample.

The data acquisition device evaluates the acquired data to obtain two parallel immune response detection results, which are respectively calculated as a first measured value and a second measured value, and transmits the measured values to the processor.

The processor is capable of calculating the ratio of the first measured value to the second measured value of the sample to be detected, comparing the ratio with the stored critical point c, and further identifying whether the sample to be detected is a HOOK effect sample.

In some embodiments of the invention, the reaction curve a, the reaction curve B and the critical point c stored in the processor are obtained by performing a method comprising the steps of:

a1, detecting a series of known standard substances with different concentrations containing target molecules to be detected, wherein two parallel immunoreaction detection is carried out on the standard substances with the same target molecule concentration to be detected, and the detection results of the two parallel immunoreactions are recorded and respectively calculated as a measured value a and a measured value a'; the detection mode of the measured value a is the same as that of a first measured value of the sample to be detected, and the detection mode of the measured value a' is the same as that of a second measured value of the sample to be detected;

The third aspect of the invention relates to the use of a method according to the first aspect of the invention or a system according to the second aspect of the invention in chemiluminescent immunoassay, enzyme linked immunoassay and immunonephelometric immunoassay; preferably in chemiluminescent immunoassay; further preferred is the use in photo-activated chemiluminescence immunoassay.

Examples

In order that the invention may be more readily understood, the invention will be further described in more detail by the following examples, which are given by way of illustration only and are not limiting in scope of application. The starting materials or components used in the present invention may be prepared by commercial or conventional methods unless specifically indicated.

Alpha Fetoprotein (AFP) is a glycoprotein, also known as fetal alpha globulin, belonging to the albumin family. The AFP (alpha fetoprotein) value in serum of a Primary Liver Cancer (PLC) patient is greatly different, and the normal value and the pathological value are extremely bad and can reach 7 orders of magnitude. It has been reported in the literature that the concentration of AFP in primary serum of a primary liver cancer patient is directly measured by IEMA to be 29ng/mL, however, the actual concentration of AFP is measured and calculated to be 5.9X10 after serial dilution of serum samples ⁶ ng/mL. As can be seen, there are still significant drawbacks to the detection of AFP in the conventional art.

Example 1: detection of AFP samples by conventional methods

The kit adopted is an Alpha Fetoprotein (AFP) detection kit (chemiluminescence method) produced by Komebo positive diagnostic technique (Shanghai) limited company (batch number: L2001), and the main components are:

reagent 1: luminescent particles coated with AFP antibodies;

reagent 2: biotin-labeled AFP antibodies.

The testing method comprises the following steps:

1. adding 25 μl of the sample to be tested, 25 μl of the reagent 1, 25 μl of the reagent 2 into the reaction well, and incubating at 37deg.C for 15min;

2. to the reaction well, 175. Mu.l of a universal solution for a photo-activated chemiluminescent assay system (donor reagent) was added, and incubated at 37℃for 10min, using

The analyzer takes readings.

Test sample (collected from clinical serum samples):

sample 1: negative serum sample (true measurement about 5 ng/mL)

Sample 2: low positive serum samples (true assay about 100 ng/mL)

Sample 3: strong positive serum samples (true assay about 2X 10) ⁶ ng/mL)

The test results are shown in Table 1.

TABLE 1

Initial measurement value	Measured value ng/mL
		Sample 1	7.41
Sample 2	109.43
		Sample 3	11.79

The results shown in Table 1 are the results of direct detection by the conventional method, the measured value of sample 3 is 11.79ng/mL, and if the clinical manifestation is not combined, the sample is easily misjudged as a weak positive sample, and the sample cannot be identified as a HOOK effect sample.

In the case where sample 3 is known to be a strong positive sample, it is re-detected after 50-fold dilution with a diluent, and the detection results are shown in table 2.

TABLE 2

50 times dilution	Measured value ng/mL
		Diluting the sample	>1000

As above, sample 3 after 50-fold dilution was >1000ng/mL, which was determined to be a HOOK sample.

Example 2: identification of HOOK Effect samples Using the methods of the invention

Testing standard substances: the concentration range is 0 ng/mL-4X 10 ⁶ ng/mL of purified AFP antigen solution

Test sample (collected from clinical serum samples):

sample 1: negative (true measured value about 5 ng/mL)

Sample 2: positive with low value (true measured value about 100 ng/mL)

Sample 3: strong positives (true measures about 2X 10 ⁶ ng/mL)

The main components of the adopted kit are as follows:

reagent 1: luminescent microparticles coated with AFP antibody (concentration 100. Mu.g/mL), biotin-labeled AFP antibody (concentration 2. Mu.g/mL);

reagent 2: luminescent microparticles coated with AFP antibody (concentration 20. Mu.g/mL), biotin-labeled AFP antibody (concentration 0.4. Mu.g/mL).

The testing method comprises the following steps:

the two reaction vessels are the same sample test group, and the following liquid adding steps 1 and 2 are repeated for different sample test groups:

1. adding 10 μl of the sample to be tested and 25 μl of the reagent 1 into the reaction well 1;

2. adding 10 μl of the sample to be tested and 25 μl of the reagent 2 into the reaction well 2;

3. incubating each reaction well at 37 ℃ for 15min;

4. to each reaction well, 175. Mu.l of a universal solution (donor reagent) for a photo-activated chemiluminescent assay system was added, and incubated at 37℃for 10min, using

The analyzer takes readings.

The test results of the gradient diluted AFP standards (Nos. 1-20) were shown in Table 3.

A reaction curve A and a reaction curve B of the standard substance concentration and the signals of the reagent 1 and the reagent 2 are respectively prepared according to the numerical values of the table 3, as shown in figure 3; it can be seen that the front zone region of the reaction curve a corresponds to the standard substance 1-11, and the rear zone region of the reaction curve B corresponds to the standard substance 9-20, so that the portion where the front zone region of the reaction curve a overlaps with the concentration existing in the rear zone region of the reaction curve B is the concentration range of the standard substance 9-11, the corresponding a/B signal ratio is 15.05-24.31, and the central point a/B signal ratio=19.18 is taken as the critical point.

Storing the standard substance test result into

An analyzer.

Three sets of samples were tested according to the test methods described above, and the test results are shown in table 4.

TABLE 3 Table 3

TABLE 4 Table 4

Analysis of test results:

the ratio (7.87) of the first measured value to the second measured value of the sample 1 is smaller than the critical point c (19.18), and the ratio (11.18) of the first measured value to the second measured value of the sample 2 is smaller than the critical point c (19.18), so that the sample 1 and the sample 2 are both normal samples (non-HOOK effect samples);

the first value/second value ratio (339.41) of sample 3 is greater than the critical point c (19.18), and the first value (427658) is less than the value a (7932206), so that sample 3 is a HOOK effect sample.

Therefore, the method can directly, simply, conveniently, quickly and accurately judge whether the sample to be detected is the HOOK effect sample, thereby avoiding missed detection or detection errors caused by the HOOK effect.

Example 4: system for identifying HOOK effect sample

The main components of the adopted kit are as follows:

The system comprises:

two reaction cups which are centrally symmetrical on the immune reaction device are set to be the same sample test group, and the measured value with larger ratio of the content of the specific capture molecules in the detection reagent to the content of the target molecules to be detected in the sample to be detected is the first measured value, and the other is the second measured value.

The detection steps executed by the system are as follows:

1. the controller transfers two reaction cups filled with the same sample to be tested and corresponding detection reagent in the immune reaction device to the immune determination device;

2. the light exciter emits laser and irradiates the laser into the reaction cup, so that a compound formed by target molecules to be detected in the sample to be detected and specific capture molecules in the detection reagent is excited to emit light signals;

3. the counter receives the optical signals and simultaneously performs data acquisition on two parallel immune response detections, and records that the detection results of the immune response are a first measurement value and a second measurement value respectively;

4. the processor calculates the ratio of the first measured value to the second measured value of the same sample to be detected, and retrieves the stored data to judge whether the sample is the HOOK effect sample.

The results of the detection are shown in tables 5 and 6 below.

Table 5: measurement result of critical point c of standard substance

Table 6: measurement result of sample to be measured

Analysis of results: the ratio of the first measured value to the second measured value of the sample 03 and the sample 08 is smaller than the critical point c (15.18), and the sample is a normal sample (a sample with no HOOK effect); the ratio of the first measurement value to the second measurement value of the sample 11 is greater than the critical point c (15.18), and the first measurement value (7974192) is greater than the measurement value a (6848780), and is also a normal sample (a non-HOOK effect sample); the first value/second value ratio of sample 15 is greater than the critical point c (15.18), and the first value (4321968) is less than the value a (6848780), which is a HOOK effect sample.

It should be noted that the above-described embodiments are only for explaining the present invention and do not constitute any limitation of the present invention. The invention has been described with reference to exemplary embodiments, but it is understood that the words which have been used are words of description and illustration, rather than words of limitation. Modifications may be made to the invention as defined in the appended claims, and the invention may be modified without departing from the scope and spirit of the invention. Although the invention is described herein with reference to particular means, materials and embodiments, the invention is not intended to be limited to the particulars disclosed herein, as the invention extends to all other means and applications which perform the same function.

Claims

1. A method of identifying a sample of the HOOK effect in an immunoassay comprising the steps of:

s3, acquiring a critical point c of a standard substance, wherein detection results of two parallel immune reactions of the standard substance corresponding to the critical point c are respectively calculated as a measured value a and a measured value a ', the measured value a and a first measured value of the sample to be detected are detected in the same mode, and the measured value a' and a second measured value of the sample to be detected are detected in the same mode;

2. The method according to claim 1, wherein the sample to be tested is a non-HOOK effect sample when the ratio of the first measurement value to the second measurement value is smaller than the critical point c;

3. The method of claim 1, wherein the sample to be tested is a HOOK effect sample when the ratio of the first measurement value to the second measurement value is greater than the critical point c and the first measurement value is less than the measurement value a.

4. A method according to any one of claims 1-3, wherein the ratio of the content of target molecules to be detected/the content of specific capture molecules in the two parallel immunoreactions assays is different; wherein the specific capture molecule is capable of specifically binding to the target molecule to be detected.

5. The method according to claim 4, wherein the two parallel immunoreactions are detected with a first or a and a second or a' of the immunoreactions having a greater ratio of the amount of specific capture molecule to the amount of target molecule to be detected.

6. The method according to any one of claims 1-5, wherein in step S2, the method of obtaining the critical point c comprises the steps of:

7. The method according to any one of claims 1 to 6, wherein the target molecule to be tested is selected from an antigen or an antibody; and/or

8. The method of claim 7, wherein the content of the specific capture molecule is the content of the first capture molecule, the content of the second capture molecule, or the sum of the content of the first capture molecule and the content of the second capture molecule; preferably the content of the first capture molecules.

9. A system for implementing the method of any one of claims 1-8, comprising:

the immunoassay device is provided with a data acquisition device;

10. The system of claim 9, wherein the two reaction cups for carrying the same sample to be tested and corresponding test reagents are placed at any given location on the immunoreaction apparatus; preferably, the two reaction cups for carrying the same sample to be tested and the corresponding detection reagents are positioned adjacent to each other on the immunoreaction device.

11. The system according to claim 9 or 10, wherein the immunoassay device performs data acquisition on two parallel immunoreaction assays of the same sample to be tested; preferably, the immunoassay device performs data acquisition on two parallel immunoreaction detections of the same sample to be tested at the same time.

12. The system according to any one of claims 9-11, wherein the processor has stored therein a response curve a, a response curve B and a critical point c for identifying whether the sample to be tested is a HOOK effect sample.

13. Use of the method according to any one of claims 1 to 8 or the system according to any one of claims 9 to 12 in chemiluminescent immunoassay, enzyme linked immunoassay and immunonephelometric immunoassay; preferably in chemiluminescent immunoassay; further preferred is the use in photo-activated chemiluminescence immunoassay.