Composition for cell lysis as well as application and kit thereof

Abstract

The invention provides a composition for cell lysis as well as application and a kit thereof, and relates to the technical field of biology. The composition is prepared from the following components in parts by mass: 0.5 to 5 parts of serum albumin, 1 to 10 parts of Triton X-100, 0.1 to 1 part of ethylene diamine tetraacetic acid salt and 0.05 to 0.2 part of a surfactant S-22. The composition can effectively lyse epithelial cells and improve the subsequent detection effect on analyzed substances in epithelial cell lysis products.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a composition for cell lysis and an application and a kit thereof. Background Art

[0002] The following statements merely provide background information related to the present disclosure and do not necessarily constitute prior art.

[0003] Molecular biological detection methods are helpful for rapid detection of pathogens of respiratory diseases. Most of the detection objects of molecular biological detection methods involve the detection of cell lysis products, such as proteins, DNA and RNA in cells or on cell membranes, so cells need to be lysed in the pretreatment stage of most detection methods. Cell lysis solution is a solution used to destroy cell membranes and nuclear membranes and extract proteins and other molecules in cells. The function of cell lysis solution significantly affects the test results.

[0004] In in vitro diagnosis, most diagnostic samples include epithelial cells, such as throat swabs and nasal swabs. Although there are currently a variety of cell lysates, such as RIPA lysate and IP cell lysate, the difference between different lysates in the prior art mainly lies in the difference in the intensity of cell or tissue lysis, but there is a lack of cell lysates for a specific cell type, such as epithelial cells. Sufficient cell lysis helps to obtain more target molecules during the detection process and improve the detection effect. Therefore, there is an urgent need for a cell lysate that can effectively lyse epithelial cells.

[0005] In view of this, the present invention is proposed. Summary of the invention

[0006] The object of the present invention is to provide a composition for cell lysis and application thereof, so as to improve the effect of epithelial cell lysis.

[0007] In order to solve the above technical problems, the present invention particularly adopts the following technical solutions:

[0008] In a first aspect, a composition for cell lysis is provided, which comprises, by weight, 0.5 to 5 parts of serum albumin, 1 to 10 parts of Triton X-100, 0.1 to 1 part of ethylenediaminetetraacetate, and 0.05 to 0.2 parts of surfactant S-22.

[0009] In a second aspect, there is provided use of the composition for cell lysis of the first aspect in lysing epithelial cells for non-diagnostic and non-therapeutic purposes, or in preparing a kit for detecting epithelial cell lysis products.

[0010] In a third aspect, a detection kit is provided, which comprises the composition for cell lysis according to the first aspect.

[0011] Compared with the prior art, the present invention has the following beneficial effects:

[0012] The composition for cell lysis provided by the present invention comprises serum albumin, Triton X-100, ethylenediaminetetraacetate and surfactant S-22. The chemicals used in the composition are easy to obtain, simple in configuration, and without complex processes. The technical solution is relatively easy to implement. The composition helps to rupture epithelial cells, so that the contents of epithelial cells are more fully released into the system to be detected, and the content and concentration of the target to be detected in the fixed volume detection system are increased, so that the target to be detected is more fully identified by the detection reagent. The composition also helps to release housekeeping proteins in epithelial cells and improve the signal value of the internal reference when the housekeeping proteins are used as internal references.

[0013] In a preferred embodiment, the target is detected by immunochromatography technology, and the target to be detected can be retained as much as possible by the antibody coated on the immunochromatography test paper during the immunochromatography process, thereby improving the reading signal and thus improving the detection sensitivity. At the same time, the composition can effectively eliminate the background of the immunochromatography test paper, improve the chromatographic membrane surface, and improve the reading to make the result clearer and easier to obtain. BRIEF DESCRIPTION OF THE DRAWINGS

[0014] In order to more clearly illustrate the specific implementation of the present invention or the technical solution in the prior art, the following will briefly introduce the drawings required for the specific implementation or the prior art description. Obviously, the drawings described below are some implementations of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work. (The actual experiment is an average of three parallel samples, but the attached drawings only provide one sample for a clearer membrane surface comparison)

[0015] Figure 1 This is a photo of the test strips after the samples were lysed using lysing solution 3 and lysing solution 4 in Example 2 and then tested using the new coronavirus latex immunochromatographic test paper, with lysing solution 3 on the left and lysing solution 4 on the right;

[0016] Figure 2 These are photos of the test strips after the samples were lysed using lysis solution 3 and lysis solution 4 respectively in Example 2 and then detected using the novel coronavirus fluorescent immunochromatographic test strips for excitation and fluorescence color development, with lysis solution 4 on the left and lysis solution 3 on the right. DETAILED DESCRIPTION

[0017] The technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments. Obviously, the described embodiments are part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0018] As used herein, "prepared from" is synonymous with "comprising". As used herein, the terms "comprising", "including", "having", "containing" or any other variation thereof, are intended to cover a non-exclusive inclusion. For example, a composition, process, method, article, or apparatus that comprises the listed elements is not necessarily limited to only those elements but may include other elements not expressly listed or inherent to such composition, process, method, article, or apparatus.

[0019] As used herein, the conjunction "consisting of excludes any unspecified elements, steps or components. If used in a claim, this phrase will render the claim closed such that it excludes materials other than those described, except for normal impurities associated therewith.

[0020] Herein, when an amount, concentration, or other value or parameter is expressed as a range, a preferred range, or a range defined by a series of upper preferred values ​​and lower preferred values, this should be understood to specifically disclose all ranges formed by any pairing of any range upper limit or preferred value with any range lower limit or preferred value, regardless of whether the range is disclosed separately. For example, when a range of "1 to 5" is disclosed, the described range should be interpreted as including ranges "1 to 4", "1 to 3", "1 to 2", "1 to 2 and 4 to 5", "1 to 3 and 5", etc. When a numerical range is described herein, unless otherwise stated, the range is intended to include its end values ​​and all integers and fractions within the range.

[0021] As used herein, "and / or" is used to indicate that one or both of the described situations may occur, for example, A and / or B includes (A and B) and (A or B).

[0022] Herein, unless otherwise indicated, arbitrary numbering is used to distinguish one entity or action from another entity or action, and does not necessarily require or imply any actual such relationship, order, or importance between such entities or actions.

[0023] As used herein, unless otherwise stated, "optionally", "optional", "optional" or "optional" means that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs or does not occur.

[0024] In a first aspect, a composition for cell lysis is provided, which comprises serum albumin, TritonX-100, ethylenediaminetetraacetate and surfactant S-22.

[0025] Serum albumin can act as a stabilizer to enhance the stability of cell lysis products; it can also simulate physiological environments and adjust the osmotic pressure of the composition. The composition includes 0.5 to 5 parts of serum albumin by weight, for example, but not limited to 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5 or 5 parts.

[0026] In an alternative embodiment, the serum albumin comprises bovine serum albumin (BSA).

[0027] Triton X-100 is a non-ionic detergent with a chemical name of octylphenoxypolyethoxyethanol. Triton X-100 can destroy the lipid bilayer structure of the cell membrane, release the cell contents, and dissociate membrane proteins and other protein complexes. In terms of mass fraction, the composition includes 1 to 10 parts of Triton X-100, for example, but not limited to 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 parts.

[0028] Ethylenediaminetetraacetate (EDTA) can form stable complexes with various metal ions, is a commonly used chelating agent, has strong complexing ability, and can form stable water-soluble complexes with various metal ions, thereby removing or inhibiting the activity of these metal ions. In terms of mass fractions, the composition includes 0.1 to 1 part of ethylenediaminetetraacetate, for example, but not limited to 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1 part.

[0029] In an optional embodiment, the ethylenediaminetetraacetic acid salt includes one or more of a metal salt and an ammonium salt. Metal salts include, but are not limited to, salts of alkali metals, alkaline earth metals, and transition metals, and exemplary metal salts include potassium salts, sodium salts, magnesium salts, calcium salts, or zinc salts. In an optional embodiment, the ethylenediaminetetraacetic acid salt includes tetrasodium ethylenediaminetetraacetic acid.

[0030] The molecular formula of surfactant S-22 is C 30 H 62 O 10 , is a nonionic surfactant that can promote protein dissolution. Surfactant S-22 also has a wetting and penetrating effect, which is beneficial to the immunochromatography process. According to the mass fraction, the composition includes 0.05 to 0.2 parts of surfactant S-22, for example, but not limited to 0.05, 0.1, 0.15 or 0.2 parts.

[0031] In an optional embodiment, the composition for cell lysis also includes NP-40 (Phenoxypolyethoxyethanol), which is a non-ionic polyoxyethylene surfactant that can dissolve lipids on the cell membrane, thereby destroying the cell membrane structure and releasing proteins and other molecules inside the cell.

[0032] In an optional embodiment, the composition includes 0.5 to 2 parts of NP-40 by weight, for example, but not limited to, 0.5, 0.6, 0.8, 0.9, 1.0, 1.2, 1.5, 1.8 or 2 parts.

[0033] In an optional embodiment, the composition further comprises a buffer component and / or a preservative.

[0034] "Buffer component" refers to a component in an aqueous solution or composition that enables the aqueous solution or composition to resist and / or inhibit changes in pH when an acid or base is added to the solution or composition. The buffer component can be selected from a buffer salt or a non-ionic buffer component. In an optional embodiment, the buffer component includes tris (hydroxymethyl) aminomethane (Tris), preferably including 0.1 to 1 parts of tris (hydroxymethyl) aminomethane, for example, but not limited to 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1 part.

[0035] In an optional embodiment, the preservative includes sodium azide, preferably 0.05-0.3 parts of sodium azide, for example, but not limited to 0.05, 0.1, 0.15, 0.2, 0.25 or 0.3 parts.

[0036] In an optional embodiment, the composition further comprises NaCl, preferably 4 to 5 parts of NaCl, for example, but not limited to 4, 4.5 or 5 parts.

[0037] In an optional embodiment, the composition includes, by weight: 4 to 5 parts of NaCl, 0.5 to 5 parts of bovine serum albumin, 1 to 10 parts of Triton X-100, 0.1 to 1 part of tetrasodium ethylenediaminetetraacetate, 0.05 to 0.3 parts of sodium azide, 0.1 to 1 part of tris(hydroxymethyl)aminomethane, 0.05 to 0.2 parts of surfactant S-22 and 0.5 to 2 parts of NP-40.

[0038] In an optional embodiment, the composition includes, by weight: 4 parts of NaCl, 1 part of bovine serum albumin, 5 parts of Triton X-100, 0.5 parts of tetrasodium ethylenediaminetetraacetate, 0.2 parts of sodium azide, 0.6 parts of tris(hydroxymethyl)aminomethane, 0.1 parts of surfactant S-22 and 1 part of NP-40.

[0039] In an optional embodiment, the composition further comprises a solvent, which includes but is not limited to water, a buffer solution or a non-buffer solution.

[0040] In an optional embodiment, the composition further comprises a solvent, and the composition is a cell lysate; the concentrations of the components in the composition are: NaCl 4-5% wt, for example, it can be but not limited to 4, 4.5 or 5% wt; bovine serum albumin 0.5-5% wt, for example, it can be but not limited to 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5 or 5% wt; Triton X-100 1-10%wt, for example, but not limited to 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10%wt; 0.1-1%wt of tetrasodium ethylenediaminetetraacetate, for example, but not limited to 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1%wt; 0.05-0.3%wt of sodium azide, for example, but not limited to 0.05, 0.1, 0.15, 0.2, 0.25 or 0.3%wt; 0.1-1%wt of tris(hydroxymethyl)aminomethane, for example, but not limited to 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1%wt; surfactant S-22 0.05-0.2%wt, for example, but not limited to 0.05, 0.1, 0.15 or 0.2%wt; and, NP-40 0.5-2%wt, for example, but not limited to 0.05, 0.1, 0.2, 0.3, 0.5, 0.6, 0.8, 0.9, 1.0, 1.2, 1.5, 1.8 or 2%wt. In this embodiment, the solvent may optionally be water.

[0041] In an optional embodiment, the composition further comprises a solvent, and the composition is a cell lysate; the concentrations of the components in the composition are: NaCl 4%wt, serum albumin 1%wt, Triton X-100 5%wt, tetrasodium ethylenediaminetetraacetate 0.5%wt, sodium azide 0.2%wt, tris(hydroxymethyl)aminomethane 0.6%wt, surfactant S-22 0.1%wt and NP-40 1%wt. In this embodiment, the solvent is optionally water.

[0042] In a second aspect, there is provided a use of the composition for cell lysis described in the first aspect in lysing epithelial cells for non-diagnostic and therapeutic purposes, or in preparing a kit for detecting epithelial cell lysis products.

[0043] In an optional embodiment, the detecting of epithelial cell lysate includes detecting an internal reference protein.

[0044] In an optional embodiment, the internal reference protein includes but is not limited to one or more of glyconuclease P, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), β-microglobulin, β-tubulin, α-tubulin, 18sRNA, β-actin and TATA binding protein.

[0045] In a third aspect, a detection kit is also provided, which comprises the composition for cell lysis described in the first aspect, and can realize the detection of cell lysis products.

[0046] In an optional embodiment, the detection kit further comprises a reagent for detecting cell lysis products.

[0047] In an optional embodiment, the detection kit also includes a detection reagent for any of the following detection methods: immunofluorescence staining detection, flow sorting detection, immunoblotting detection, immunohistochemistry detection, ELISA detection, immunochromatography detection, immunoprecipitation detection, immunocoprecipitation detection or immunomagnetic bead detection.

[0048] In an optional embodiment, the detection kit further comprises a reagent for detecting an internal reference protein.

[0049] In an optional embodiment, the internal reference protein includes one or more of glyconuclease P, glyceraldehyde-3-phosphate dehydrogenase, β-microglobulin, β-tubulin, α-tubulin, 18sRNA, β-actin and TATA binding protein.

[0050] In an optional embodiment, the detection kit includes immunochromatographic test paper.

[0051] In an optional embodiment, the quality control line of the immunochromatographic test paper uses the internal reference protein as the detection target molecule.

[0052] In an optional embodiment, the marker of the immunochromatographic test paper includes, but is not limited to, one or more of fluorescent particles, colored particles, latex particles, quantum dots, chemiluminescent reagents, fluorescent molecules, colloidal gold, colloidal silver, colloidal selenium, horseradish peroxidase, and alkaline phosphatase, preferably fluorescent particles, latex particles, or colloidal gold. The composition for cell lysis provided in the first aspect can effectively eliminate the background of the immunochromatographic test paper using fluorescent particles and latex particles, improve the chromatographic membrane surface, and improve the reading to make the result clearer and easier to obtain.

[0053] The present invention is further described below by means of specific examples. However, it should be understood that these examples are only used for more detailed description and should not be construed as limiting the present invention in any form.

[0054] Example 1

[0055] Prepare the following lysis buffer according to Table 1:

[0056] Table 1

[0057] Ingredients\Concentration (%wt) Lysis buffer 1 Lysis buffer 2 Lysate 3 NaCl 4 4 4 BSA 1 1 1 Triton X-100 5 5 5 Tetrasodium EDTA 0.5 0.5 0.5 Sodium Azide 0.2 0.2 0.2 Tris(hydroxymethyl)aminomethane - 0.6 0.6 Surfactant S-22 - - 0.1

[0058] Collect 50 human nasal swab negative sample cotton swabs, put each nasal swab cotton swab into a lysis tube, add 400 μL of normal saline and soak for at least 5 minutes, use the thumb and index finger to squeeze the lysis tube 20 times, draw out the cotton swab to get the nasal swab eluate, mix the 50 nasal swab eluates into a homogenized sample solution as the negative matrix of the sample, use the lysis solution in Table 1 for influenza A virus latex immunochromatography test paper for detection, take the homogenized negative sample solution and mix it with the lysis solution in Table 1 respectively, the mixing volume ratio is 1:1, let it stand for more than 15 minutes after mixing, and then load the sample, the loading volume is 100 μL, and 3 or more parallel samples are used for testing, and the color value or average value of more than 95% of the parallel samples is taken as the result.

[0059] Interpretation of latex immunochromatography test reagent results: The color values ​​of the test line and the quality control line refer to a colorimetric card disclosed in the invention patent with publication number CN111909257A (application number CN202010736865.X, invention name "A method for synthesizing carlipodol antigen and its application"). The higher the number, the darker the color. The color value range is 1-10, and the chromaticity of the reading of 3.5 and above is determined to be visible to the naked eye. The depth of the C line and T line in the subsequent tables are expressed in this reading. Therefore, the criterion for the visible line is whether the color chromaticity of the C line and T line reaches the color card reading of 3.5, where "-" indicates negative.

[0060] The experimental results are shown in the following table:

[0061] Table 2

[0062] Lysis buffer C line output reading Lysis buffer 1 7 Lysis buffer 2 7+ Lysate 3 8- PBS control group - Normal saline control group -

[0063] The results of Example 1 show that, for influenza A virus latex immunochromatographic test paper, lysis solution 3 enhances the C line signal value, indicating that the sample solution to be tested contains more GAPDH, and also indicating that the sample lysis solution 3 of Example 1 can more effectively lyse the cells in the human nasal swab and release the contents.

[0064] Example 2

[0065] Prepare the following lysis buffer according to Table 3:

[0066] Table 3

[0067] Ingredients\Concentration (%wt) Lysate 3 Lysis buffer 4 NaCl 4 4 BSA 1 1 Triton X-100 5 5 Tetrasodium EDTA 0.5 0.5 Sodium Azide 0.2 0.2 Tris(hydroxymethyl)aminomethane 0.6 0.6 Surfactant S-22 0.1 0.1 NP-40 N / A 1

[0068] The lysate in Table 3 was used for the novel coronavirus fluorescent immunochromatographic test and the novel coronavirus latex immunochromatographic test paper to detect the sample in Example 1. The sample processing method was the same as in Example 1. Three or more parallel samples were used for testing, and the results were taken as the color development value or average value of more than 95% of the parallel samples.

[0069] The interpretation of the results of the fluorescent immunochromatographic detection reagent comes from the dry immunofluorescence analysis readings of Wuxi Tianzong Yijun Biological, model: TFL1600. The reading value indicates the strength of the fluorescent signal. The larger the reading value, the stronger the fluorescent signal.

[0070] The method for interpreting the results of the latex immunochromatography detection reagent is the same as that in Example 1.

[0071] The experimental results are shown in the following table:

[0072] Table 4

[0073]

[0074] According to the results of Example 2 and Figure 1 It can be seen that when using latex immunochromatographic test paper for testing, the membrane surface of lysate 3 group was red, while the membrane surface of lysate 4 group was clean, and the C line was also clearer, indicating that in the new crown latex immunochromatographic test paper, the addition of NP-40 to the buffer weakened the matrix effect. When using fluorescent immunochromatographic test paper for testing, fluorescent particles remained on the membrane surface of lysate 3 group, while the membrane surface of lysate 4 group was clean, the reading was clearer, and the reading was improved ( Figure 2 ). The conclusion is that adding NP-40 to the buffer can reduce the matrix effect, significantly eliminate the background, increase the detection signal value, improve the membrane surface, and reduce the matrix effect to a certain extent.

[0075] Example 3

[0076] Prepare the following lysis buffer according to Table 5:

[0077] Table 5

[0078]

[0079]

[0080] The lysate in Table 5 was used for influenza A virus latex immunochromatographic test paper to detect the sample in Example 1. The sample was detected according to the method in Example 1. The results are shown in Table 6.

[0081] Table 6

[0082] Lysis buffer C line output reading Lysis buffer 5 9 Lysis buffer 6 8+ Lysis buffer 7 8+ PBS control group -

[0083] The results of Example 3 show that lysis buffers 5 to 7 can effectively lyse human epithelial cells and fully release GAPDH therein, among which lysis buffer 5 has the highest reading, indicating that lysis buffer 5 has the best lysis effect.

[0084] Example 4

[0085] Referring to Example 3 of the invention patent CN118624896A (application number CN202410857418.8, the invention name is "Immunochromatographic test paper that can ensure the effectiveness of sampling and its application in the preparation of human sample detection products" Chinese patent application), the eluate of negative oral exudate is used as the detection matrix. HIV colloidal gold immunochromatographic test paper and lysis solution 5 are used to collect the homogenized eluate of negative oral exudate as a negative sample for sample loading and detection, and the C line is observed. The following experimental groups are all tested with 3 or more parallel samples, and the experimental results are shown in Table 7 below.

[0086] Table 7

[0087] Lysis buffer C line output reading PBS 7 Normal saline 7 Lysis buffer 5 8

[0088] The results of Example 4 show that when Lysis Buffer 5 is used as a diluent for loading, the C line is stronger than that of saline and PBS as diluents. Therefore, it can be shown that in this loading system, the HSA target in the Lysis Buffer 5 loading system is more than that in PBS and saline, thus proving that the Lysis Buffer 5 formula can effectively lyse the oral epithelial cells in the negative sample and release the detection target.

[0089] Example 5

[0090] The hepatitis B surface antigen colloidal gold immunochromatographic test strip was used, and negative human whole blood was used as the negative sample for sample loading detection. The C line detection target was hemoglobin in human whole blood red blood cells. The test strip was used to load the collected negative human whole blood homogenized eluate as a negative sample for sample loading detection, and the C line was observed. The following experimental groups were tested with 3 or more parallel samples, and the experimental results are shown in Table 8:

[0091] Table 8

[0092] Lysis buffer C line output reading PBS 8 Normal saline 8 Lysis buffer 5 8

[0093] The results of Example 5 show that Lysis Buffer 5 can be used as a diluent for loading, and the C-line reading is not significantly different from the C-line with saline and PBS as diluents. Therefore, it can be shown that the hemoglobin target content in human whole blood red blood cells in the Lysis Buffer 5 loading system is not different from the human whole blood treated with PBS and saline in this loading system, thus proving that the Lysis Buffer 5 formula is more suitable for lysing oral epithelial cells in negative samples and releasing the detection target.

[0094] Example 6

[0095] The effects of different concentrations of sodium azide in the lysis solution on the sample lysis effect were compared. The lysis solution was prepared according to Table 9, and the influenza A virus latex immunochromatographic test paper was used for detection. The detection method was the same as in Example 1. The detection results are shown in Table 9.

[0096] Table 9

[0097]

[0098] According to the above experimental results, it can be seen that when the concentration of sodium azide is 0.2%wt, the sensitivity of the test strip is the highest. When the concentration of sodium azide is between 0.05%wt and 0.3%wt, the impact on the overall sensitivity of the test strip is within half a chromaticity. Its main function is solution preservation. The concentration of sodium azide is preferably 0.2%wt.

[0099] Example 7

[0100] The effects of different concentrations of surfactant S-22 in the lysis solution on the sample lysis effect were compared. The lysis solution was prepared according to Table 10, and influenza A virus latex immunochromatographic test paper was used for detection. The detection method was the same as in Example 1. The detection results are shown in Table 10.

[0101] Table 10

[0102]

[0103]

[0104] S-22 plays a wetting and penetrating role, which is beneficial to the immunochromatography process. According to the above table, when the surfactant concentration is in the range of 0.05%wt-0.1%wt, the detection sensitivity of the test strip is improved. However, when the concentration is 0.1%wt, the sensitivity of the test strip is the highest.

[0105] Example 8

[0106] The effects of different concentrations of tris(hydroxymethyl)aminomethane in the lysis solution on the sample lysis effect were compared. The lysis solution was prepared according to Table 11, and influenza A virus latex immunochromatographic test paper was used for detection. The detection method was the same as in Example 1. The detection results are shown in Table 11.

[0107] Table 11

[0108]

[0109] Tris(hydroxymethyl)aminomethane is mainly used as a stabilizer for the pH value of the solution. According to the above table, the amount of tris(hydroxymethyl)aminomethane added has a great influence on the sensitivity of the test strip, and the effect is optimal when the content of Tris is 0.6%wt.

[0110] Example 9

[0111] The effects of different concentrations of NP-40 in the lysis buffer on the sample lysis effect were compared. The lysis buffer was prepared according to Table 12, and influenza A virus latex immunochromatographic test paper was used for detection. The detection method was the same as in Example 1. The detection results are shown in Table 12.

[0112] Table 12

[0113]

[0114] According to the table above, when the addition amount of NP-40 is 0.5%wt~1%wt, the sensitivity of the reagent is improved. The improvement is more obvious when the addition amount is 1%wt.

[0115] The immunochromatographic test paper used in the above embodiment is as follows:

[0116] Influenza A virus latex immunochromatographic test strips:

[0117] The quality control line (C line) system described in CN116223796A (application number CN202310156113.X, Chinese patent application titled "Virus combined detection immunochromatographic test paper, test pen and test product") is used. The structure of the influenza A virus latex immunochromatographic test is as follows:

[0118] The latex immunochromatography test comprises a sample pad, a binding pad, a detection pad and a water absorption pad which are sequentially arranged on a fixed bottom plate along the chromatography direction of the sample to be tested.

[0119] The conjugate pad is coated with a first influenza A virus antibody (Hangzhou Xuke Biological, catalog number R1403) and a first human internal reference protein antibody (GAPDH antibody (catalog number: ET1601-4) of Hangzhou Huaan Biotechnology Co., Ltd.), and the first influenza A virus antibody and the first human internal reference protein antibody are respectively labeled with fluorescent particles; the detection pad includes a detection line T and a sampling in place quality control line C along the chromatography direction of the sample to be tested, and the detection line T is coated with a second influenza A virus antibody (Hangzhou Xuke Biological, catalog number R1401), and the sampling in place quality control line C is coated with a second human internal reference protein antibody (GAPDH antibody (catalog number: ET1601-4) of Hangzhou Huaan Biotechnology Co., Ltd.). The first influenza A virus antibody and the second influenza A virus antibody recognize influenza A virus N protein (nucleocapsid protein). The first human internal reference protein antibody and the second human internal reference protein antibody recognize glyceraldehyde-3-phosphate dehydrogenase (GAPDH).

[0120] The concentration of the second influenza A virus antibody in the test pad coating solution is 0.7 mg / mL, and the concentration of the second human internal reference protein antibody is 1 mg / mL. The test pad coating solution is a solution containing 0.9wt% NaCl, 0.13wt% Na2HPO4 and 3wt% sucrose. The spray volume of the test pad coating solution on the test line T and the sampling quality control line C is 1 μL / cm.

[0121] Novel coronavirus fluorescent immunochromatographic test paper:

[0122] The quality control line (C line) system described in CN116223796A (application number CN202310156113.X, Chinese patent application titled "Virus combined detection immunochromatographic test paper, test pen and test product") is used. The structure of the fluorescent immunochromatographic test is as follows:

[0123] The fluorescent immunochromatographic test comprises a sample pad, a binding pad, a detection pad and a water absorption pad which are sequentially arranged on a fixed bottom plate along the chromatography direction of the sample to be tested.

[0124] The conjugate pad is coated with the first novel coronavirus antibody (Hangzhou Xuke Biological, catalog number RM0032) and the first human internal reference protein antibody (GAPDH antibody (catalog number: ET1601-4) of Hangzhou Huaan Biotechnology Co., Ltd.), and the first novel coronavirus antibody and the first human internal reference protein antibody are respectively labeled with fluorescent particles; the detection pad includes a detection line T and a sampling in place quality control line C along the chromatography direction of the sample to be tested, and the detection line T is coated with the second novel coronavirus antibody (Hangzhou Xuke Biological, catalog number RM0033), and the sampling in place quality control line C is coated with the second human internal reference protein antibody (GAPDH antibody (catalog number: ET1601-4) of Hangzhou Huaan Biotechnology Co., Ltd.). The first novel coronavirus antibody and the second novel coronavirus antibody recognize the novel coronavirus N protein (nucleocapsid protein). The first human internal reference protein antibody and the second human internal reference protein antibody recognize glyceraldehyde-3-phosphate dehydrogenase (GAPDH).

[0125] The concentration of the second novel coronavirus antibody in the test pad coating solution is 0.5 mg / mL, and the concentration of the second human internal reference protein antibody is 1 mg / mL. The test pad coating solution is a solution containing 0.9wt% NaCl, 0.13wt% Na2HPO4 and 3wt% sucrose. The spray volume of the test pad coating solution on the test line T and the sampling quality control line C is 1μL / cm.

[0126] New coronavirus latex immunochromatographic test strips:

[0127] The quality control line (C line) system described in CN116223796A (application number CN202310156113.X, Chinese patent application for invention titled "Virus combined detection immunochromatographic test paper, test pen and test product") is used. The immunochromatographic test structure is as follows:

[0128] The latex immunochromatography test comprises a sample pad, a binding pad, a detection pad and a water absorption pad which are sequentially arranged on a fixed bottom plate along the chromatography direction of the sample to be tested.

[0129] The conjugate pad is coated with the first novel coronavirus antibody (Hangzhou Xuke Biological, catalog number RM0032) and the first human internal reference protein antibody (GAPDH antibody (catalog number: ET1601-4) of Hangzhou Huaan Biotechnology Co., Ltd.), and the first novel coronavirus antibody and the first human internal reference protein antibody are respectively labeled with fluorescent particles; the detection pad includes a detection line T and a sampling in place quality control line C along the chromatography direction of the sample to be tested, and the detection line T is coated with the second novel coronavirus antibody (Hangzhou Xuke Biological, catalog number RM0033), and the sampling in place quality control line C is coated with the second human internal reference protein antibody (GAPDH antibody (catalog number: ET1601-4) of Hangzhou Huaan Biotechnology Co., Ltd.). The first novel coronavirus antibody and the second novel coronavirus antibody recognize the novel coronavirus N protein (nucleocapsid protein). The first human internal reference protein antibody and the second human internal reference protein antibody recognize glyceraldehyde-3-phosphate dehydrogenase (GAPDH).

[0130] The concentration of the second novel coronavirus antibody in the test pad coating solution is 1.0 mg / mL, and the concentration of the second human internal reference protein antibody is 1 mg / mL. The test pad coating solution is a solution containing 0.9wt% NaCl, 0.13wt% Na2HPO4 and 3wt% sucrose. The spray volume of the test pad coating solution on the test line T and the sampling quality control line C is 1μL / cm.

[0131] HIV colloidal gold immunochromatographic test strips:

[0132] The colloidal gold immunochromatographic test paper is a HIV colloidal gold immunochromatographic test paper, which uses the immunochromatographic test paper recorded in Experimental Group 3 in Example 6 of CN118091132A (application number CN202410258366.2, Chinese patent application with invention name "CN202410258366.2"). The C line and the T line are respectively coated with human serum albumin specific antibodies, which are purchased from Hangzhou Xuke Biological Co., Ltd., with item numbers RM0083 and RM0084 respectively, and the other conditions are the same as the test strips provided in CN118091132A.

[0133] Hepatitis B surface antigen colloidal gold immunochromatographic test strips:

[0134] The chromatographic test paper of Example 6 in CN118091132A (application number CN202410857418.8, Chinese patent application entitled “Immunochromatographic test paper capable of ensuring sampling validity and its application in preparing human sample detection products”) was used.

[0135] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A composition for cell lysis, characterized in that Calculated by weight, the composition comprises: 0.5 to 5 parts of serum albumin, 1 to 10 parts of Triton X-100, 0.1 to 1 part of ethylenediaminetetraacetate and 0.05 to 0.2 parts of surfactant S-22.

2. The composition according to claim 1, characterized in that The composition further comprises NP-40, preferably 0.5 to 2 parts of NP-40.

3. The composition according to claim 1, characterized in that Ethylenediaminetetraacetic acid salts include metal salts or ammonium salts, preferably sodium salts; Optionally, the serum albumin comprises bovine serum albumin.

4. The composition according to claim 1, characterized in that Also includes buffer components and / or preservatives; Optionally, the buffer component includes tris(hydroxymethyl)aminomethane, preferably 0.1 to 1 part of tris(hydroxymethyl)aminomethane; Optionally, the preservative includes sodium azide, preferably 0.05 to 0.3 parts of sodium azide; Optionally, the composition further comprises NaCl, preferably 4 to 5 parts of NaCl.

5. The composition for cell lysis according to any one of claims 1 to 4, characterized in that: The composition comprises, by weight: 4 to 5 parts of NaCl, 0.5 to 5 parts of bovine serum albumin, 1 to 10 parts of Triton X-100, 0.1 to 1 part of tetrasodium ethylenediaminetetraacetate, 0.05 to 0.3 parts of sodium azide, 0.1 to 1 part of tris(hydroxymethyl)aminomethane, 0.05 to 0.2 parts of surfactant S-22 and 0.5 to 2 parts of NP-40; Optionally, the composition includes, by weight: 4 parts of NaCl, 1 part of bovine serum albumin, 5 parts of Triton X-100, 0.5 parts of tetrasodium ethylenediaminetetraacetate, 0.2 parts of sodium azide, 0.6 parts of tris(hydroxymethyl)aminomethane, 0.1 parts of surfactant S-22 and 1 part of NP-40.

6. The composition for cell lysis according to claim 5, characterized in that The composition further comprises a solvent; Optionally, the concentration of each component in the composition is: NaCl 4-5%wt, bovine serum albumin 0.5-5%wt, Triton X-100 1-10%wt, tetrasodium ethylenediaminetetraacetic acid 0.1-1%wt, sodium azide 0.05-0.3%wt, tris(hydroxymethyl)aminomethane 0.1-1%wt, surfactant S-22 0.05-0.2%wt and NP-40 0.5-2%wt; Optionally, the concentration of each component in the composition is: NaCl 4%wt, serum albumin 1%wt, Triton X-100 5%wt, tetrasodium ethylenediaminetetraacetate 0.5%wt, sodium azide 0.2%wt, tris(hydroxymethyl)aminomethane 0.6%wt, surfactant S-22 0.1%wt and NP-40 1%wt.

7. Use of the composition for cell lysis according to any one of claims 1 to 6 in lysing epithelial cells for non-diagnostic and non-therapeutic purposes, or in preparing a kit for detecting epithelial cell lysis products.

8. A detection kit, characterized in that A composition for cell lysis comprising any one of claims 1 to 6.

9. The detection kit according to claim 8, characterized in that Also included are reagents for detecting cell lysate; Optionally, the detection kit further comprises a detection reagent for any of the following detection methods: immunochromatography detection, immunofluorescence staining detection, flow sorting detection, immunoblotting detection, immunohistochemistry detection, ELISA detection, immunoprecipitation detection, immunocoprecipitation detection or immunomagnetic bead detection; Optionally, the detection kit includes immunochromatographic test paper; Optionally, the marker of the immunochromatographic test paper includes one or more of fluorescent particles, colored particles, latex particles, quantum dots, chemiluminescent reagents, fluorescent molecules, colloidal gold, colloidal silver, colloidal selenium, horseradish peroxidase and alkaline phosphatase; Optionally, the quality control line of the immunochromatographic test paper uses an internal reference protein as the detection target molecule.

10. The detection kit according to claim 9, characterized in that The detection kit also includes a reagent for detecting an internal reference protein; Optionally, the internal reference protein includes one or more of glyconuclease P, glyceraldehyde-3-phosphate dehydrogenase, β-microglobulin, β-tubulin, α-tubulin, 18sRNA, β-actin and TATA binding protein.

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