Method for reducing squamous cell carcinoma antigen detection background and post-treatment liquid

Abstract

The invention discloses a method and post-treatment fluid for reducing squamous cell carcinoma antigen detection background, and is characterized in that the method comprises the following steps: step 1, adding a sample, magnetic beads containing a coated antibody, a sample diluent and a labeled antibody labeled with acridinium ester, incubating, and executing a cleaning step; step 2, adding a post-treatment liquid, incubating, and executing a cleaning step; step 3, adding a substrate solution, entering a photometric module, and measuring the number of photons; wherein the post-treatment liquid comprises the following raw material components: a basic buffer solution, a nonionic surfactant, an alkali metal chloride, a metal ion chelate and an anionic surfactant. The method provided by the invention is simple and convenient to operate, and can improve the detection accuracy and maintain high sensitivity.

Description

Technical Field

[0001] The invention belongs to the technical field of medical in vitro diagnosis, and in particular relates to a method for reducing the background of squamous cell carcinoma antigen (SCC) detection and a post-processing liquid. Background Art

[0002] Squamous Cell Carcinoma Antigen (SCC) is a tumor marker that is often used for auxiliary diagnosis, disease monitoring and prognosis assessment of squamous cell carcinoma. However, in the actual detection process, SCC reactive determinants can naturally fall off from skin particles, saliva and other body fluids, and can easily be distributed into dust or aerosols through behaviors such as sneezing. Due to factors such as nonspecific binding, improper sample handling, and reagent contamination, the background value is often high, which will reduce the accuracy and reliability of the test, resulting in the sensitivity of the reagent not meeting the requirements, and even false positives.

[0003] During the development, production and use of the reagent, the background of the reagent is abnormally increased, the signal-to-noise ratio does not meet the standard, and there are serious risks in sensitivity and specificity due to the presence of SCC reactive determinants in the aerosol. Therefore, providing a new reaction mode is of certain significance to reduce the background and improve the signal-to-noise ratio. Summary of the invention

[0004] The present invention has found through research that by optimizing the reaction mode, the reaction is completed in one step, and after washing, an incubation dissociation step is added, which effectively reduces the background value of SCC detection, improves the signal-to-noise ratio and anti-interference ability of the detection, and thus improves the specificity and sensitivity of the reagent. Therefore, the first purpose of the present invention is to provide a method for reducing the background of squamous cell carcinoma antigen (SCC) detection. The second purpose of the present invention is to provide a post-treatment solution for reducing the background of squamous cell carcinoma antigen (SCC) detection.

[0005] To achieve the above object, the present invention adopts the following technical solution:

[0006] As a first aspect of the present invention, a method for reducing the background of squamous cell carcinoma antigen (SCC) detection comprises the following steps:

[0007] Step 1, adding sample, magnetic beads containing coated antibodies, sample diluent, labeled antibody labeled with acridinium ester, incubating, and performing a washing step;

[0008] Step 2: adding post-treatment solution, incubating, and performing a cleaning step;

[0009] Step 3: Add substrate solution, enter the photometric module, and measure the number of photons;

[0010] The raw material components of the post-treatment liquid include: basic buffer, non-ionic surfactant, alkali metal chloride, metal ion chelate and anionic surfactant.

[0011] According to the present invention, the content of the basic buffer is 20-90 mmol, and the pH value is 6.0-9.0;

[0012] The usage of non-ionic surfactant is 0.1%-0.3% of the solvent volume.

[0013] The amount of alkali metal chloride used is 0.45-0.9% by volume of the solvent;

[0014] The amount of metal ion chelate used is 0.05%-0.5% of the solvent volume;

[0015] The amount of anionic surfactant used is 0.01%-0.1% of the solvent volume.

[0016] According to the present invention, the basic buffer is at least one of PB buffer, MES buffer and Tris buffer.

[0017] Preferably, the pH value of the basic buffer is 7.0-8.5.

[0018] According to the present invention, the metal ion chelate is at least one of EDTA and EDTA derivatives.

[0019] Preferably, the amount of the metal ion chelate used is 0.1%-0.5% of the volume of the solvent.

[0020] Furthermore, the EDTA derivative is EDTA sodium salt, EDTA potassium salt or EDTA magnesium salt.

[0021] According to the present invention, the alkali metal chloride is at least one of sodium chloride and potassium chloride.

[0022] According to the present invention, the anionic surfactant is sodium dodecyl sulfate (SDS).

[0023] According to the present invention, the nonionic surfactant is at least one of Tween 20, Tween 80 and Triton-100.

[0024] As a second aspect of the present invention, a post-treatment solution for reducing the background of squamous cell carcinoma antigen (SCC) detection, the raw material components of the post-treatment solution include: a basic buffer, a non-ionic surfactant, an alkali metal chloride, a metal ion chelate and an anionic surfactant, wherein:

[0025] The content of the basic buffer is 20-90mmol, and the pH value is 6.0-9.0;

[0026] The usage of non-ionic surfactant is 0.1%-0.3% of the solvent volume.

[0027] The amount of alkali metal chloride used is 0.45-0.9% by volume of the solvent;

[0028] The amount of metal ion chelate used is 0.05%-0.5% of the solvent volume;

[0029] The amount of anionic surfactant used is 0.01%-0.1% of the solvent volume.

[0030] According to the present invention, the basic buffer is at least one of PB buffer, MES buffer and Tris buffer.

[0031] Preferably, the pH value of the basic buffer is 7.0-8.5.

[0032] According to the present invention, the metal ion chelate is at least one of EDTA and EDTA derivatives.

[0033] Preferably, the amount of the metal ion chelate used is 0.1%-0.5% of the volume of the solvent.

[0034] Furthermore, the EDTA derivative is EDTA sodium salt, EDTA potassium salt or EDTA magnesium salt.

[0035] According to the present invention, the alkali metal chloride is at least one of sodium chloride and potassium chloride.

[0036] According to the present invention, the anionic surfactant is sodium dodecyl sulfate (SDS).

[0037] According to the present invention, the nonionic surfactant is at least one of Tween 20, Tween 80 and Triton-100.

[0038] A method for reducing the background of squamous cell carcinoma antigen (SCC) detection of the present invention has the following beneficial effects:

[0039] 1. Improve detection accuracy: By modifying the reaction mode, the interference of nonspecific components on SCC detection can be effectively reduced, and the accuracy and reliability of tumor marker detection can be improved.

[0040] 2. Maintain high sensitivity: While reducing the background level, maintain high sensitivity detection of specific SCC to ensure that true tumor markers are not missed.

[0041] 3. Easy operation: The technical solution has simple steps, does not require complex equipment, and is easy to promote and apply in clinical laboratories.

[0042] The post-treatment liquid for reducing the background of squamous cell carcinoma antigen (SCC) detection of the present invention has the following effects: when it can be used for squamous cell carcinoma antigen (SCC) detection, it can effectively reduce the background value of SCC detection. DETAILED DESCRIPTION

[0043] The present invention is further described below in conjunction with specific examples. It should be understood that the following examples are only used to illustrate the present invention and are not used to limit the scope of the present invention.

[0044] The instrument used in the present invention is Pinfeng KunLun i2000 (Guangdong Medical Instrument No. 20242220019), which is matched with substrate liquid (Guangdong Shenzhen Medical Instrument No. 20220315) and cleaning liquid (Guangdong Shenzhen Medical Instrument No. 20211542).

[0045] Example 1

[0046] 1. Sample pretreatment

[0047] Use a vacuum blood collection tube containing separation gel to draw 5 mL of venous blood on an empty stomach according to sterile requirements. Centrifuge at 5000 r / min for 5 minutes. When using serum, operators must wear masks, head covers, and disposable latex gloves to avoid operating in the air vents. Use a pipette to take the upper serum sample, divide it into packages, and store it in the freezer.

[0048] 2. Design responsiveness:

[0049] The control group used double antibody sandwich method and one-step method.

[0050] The experimental group of the present invention adopts a double antibody sandwich method, a two-step method, in which all reactions are completed in the first step, and a special post-treatment solution is added in the second step for incubation to dissociate the unstable SCC conjugate.

[0051] 3. Put the normally coated magnetic beads, markers, and samples into the box, and add specific post-processing liquid to the experimental group. Store the reagents at room temperature and at the air outlet to prevent excessive volatilization, and cover with a semi-closed silicone plug.

[0052] The reaction mode and the formula of the post-treatment solution are shown in Tables 1 and 2.

[0053] Comparative test:

[0054] The low-value reactivity changes and overall trends were evaluated at 0 days, 3 days, 7 days, 10 days, and 14 days. The experimental results are shown in Tables 3-5.

[0055] Note: In the post-treatment solution formulas listed in Tables 1 and 2, the percentages of Triton X-100, NaCl, EDTA, SDS, KCl, Tween20 and other ingredients all represent their respective proportions in the volume of the solvent. For example, 0.9% NaCl in Table 1 means that the amount of NaCl used accounts for 0.9% of the volume of the solvent.

[0056] Table 1 Reaction mode and post-treatment liquid formulation of comparative example 1 and embodiment 1

[0057]

[0058] Table 2 Reaction mode and post-treatment liquid formulation of Examples 1-3

[0059]

[0060]

[0061] Table 3 Changes and overall trends of low-value reactivity of comparative example 1

[0062]

[0063] Table 4 Low value reactivity changes and overall trends of Example 1

[0064]

[0065] Table 5 Low value reactivity changes and overall trends of Example 2

[0066]

[0067] Table 6 Low value reactivity changes and overall trends of Example 3

[0068]

[0069] Table 7 Low value reactivity changes and overall trends of Example 4

[0070]

[0071] According to the analysis of the results on day 0, the background of Examples 1-4 with modified reaction modes was lower than that of the control group, and the reaction gradient was significantly better than that of the control group. Examples 1-4 showed good sensitivity in the low value range.

[0072]

[0073] Through the data analysis of the control group and the experimental group from 0 days to 14 days, under the same conditions, the daily precision CV of the experimental group was more stable and the ability to resist aerosol pollution was stronger. The background of the control group increased by 4 times during the change from 0 days to 14 days. It can be seen that after changing the reaction mode, adding post-processing liquid can improve the anti-pollution ability of the reagent to a certain extent.

[0074] In summary, the present invention provides a method for reducing the background of squamous cell carcinoma antigen (SCC) detection. By optimizing the reaction mode, the reaction is completed in one step. After washing, an incubation dissociation step is added, which effectively reduces the background value of SCC detection, improves the signal-to-noise ratio and anti-interference ability of the detection, and thus improves the specificity and sensitivity of the reagent.

[0075] The specific embodiments of the present invention are described in detail above, but they are only examples, and the present invention is not limited to the specific embodiments described above. For those skilled in the art, any equivalent modifications and substitutions made to the invention are also within the scope of the present invention. Therefore, the equalization changes and modifications made without departing from the spirit and scope of the present invention should be included in the scope of the present invention.

Claims

1. A method for reducing the background of squamous cell carcinoma antigen detection, characterized in that: The steps include: Step 1, adding sample, magnetic beads containing coated antibodies, sample diluent, labeled antibody labeled with acridinium ester, incubating, and performing a washing step; Step 2: adding post-treatment solution, incubating, and performing a cleaning step; Step 3: Add substrate solution, enter the photometric module, and measure the number of photons; The raw material components of the post-treatment liquid include: basic buffer, non-ionic surfactant, alkali metal chloride, metal ion chelate and anionic surfactant.

2. The method for reducing squamous cell carcinoma antigen detection background according to claim 1, characterized in that: The content of the basic buffer is 20-90 mmol, and the pH value is 6.0-9.0; The usage of non-ionic surfactant is 0.1%-0.3% of the solvent volume. The amount of alkali metal chloride used is 0.45-0.9% by volume of the solvent; The amount of metal ion chelate used is 0.05%-0.5% of the solvent volume; The amount of anionic surfactant used is 0.01%-0.1% of the solvent volume.

3. The method for reducing squamous cell carcinoma antigen detection background according to claim 1 or 2, characterized in that: The basic buffer is at least one of PB buffer, MES buffer and Tris buffer.

4. The method for reducing squamous cell carcinoma antigen detection background according to claim 1 or 2, characterized in that: The metal ion chelate is at least one of EDTA and EDTA derivatives; and / or, The EDTA derivative is EDTA sodium salt, EDTA potassium salt or EDTA magnesium salt; and / or, The alkali metal chloride is at least one of sodium chloride and potassium chloride; and / or, The anionic surfactant is sodium lauryl sulfate; and / or, The nonionic surfactant is at least one of Tween 20, Tween 80 and Triton-100.

5. The method for reducing the background of squamous cell carcinoma antigen detection according to claim 1, 2 or 3, characterized in that: The amount of the metal ion chelate used is 0.1%-0.5% of the volume of the solvent.

6. The method for reducing the background of squamous cell carcinoma antigen detection according to claim 1, 2 or 3, characterized in that: The EDTA derivative is EDTA sodium salt, EDTA potassium salt or EDTA magnesium salt.

7. A post-treatment solution for reducing the background of squamous cell carcinoma antigen detection, characterized in that: The raw material components of the post-treatment liquid include: basic buffer, non-ionic surfactant, alkali metal chloride, metal ion chelate and anionic surfactant, wherein: The content of the basic buffer is 20-90mmol, and the pH value is 6.0-9.0; The usage of non-ionic surfactant is 0.1%-0.3% of the solvent volume. The amount of alkali metal chloride used is 0.45-0.9% by volume of the solvent; The amount of metal ion chelate used is 0.05%-0.5% of the solvent volume; The amount of anionic surfactant used is 0.01%-0.1% of the solvent volume.

8. The post-treatment solution for reducing squamous cell carcinoma antigen detection background according to claim 7, characterized in that: The metal ion chelate is at least one of EDTA and EDTA derivatives; and / or, The EDTA derivative is EDTA sodium salt, EDTA potassium salt or EDTA magnesium salt; and / or, The alkali metal chloride is at least one of sodium chloride and potassium chloride; and / or, The anionic surfactant is sodium lauryl sulfate; and / or, The nonionic surfactant is at least one of Tween 20, Tween 80 and Triton-100.

9. The post-treatment solution for reducing the background of squamous cell carcinoma antigen detection according to claim 7 or 8, characterized in that: The amount of the metal ion chelate used is 0.1%-0.5% of the volume of the solvent.

10. The post-treatment solution for reducing the background of squamous cell carcinoma antigen detection according to claim 7 or 8, characterized in that: The EDTA derivative is EDTA sodium salt, EDTA potassium salt or EDTA magnesium salt.

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