Human immunodeficiency virus (HIV) antibody and application and detection product thereof

Abstract

The invention provides a pair of human immunodeficiency virus (HIV) antibodies as well as application and a detection product thereof. The pair of human immunodeficiency virus HIV antibodies provided by the invention comprises an antibody c and / or an antibody n, and provides amino acid sequences of a heavy chain complementary determining region and a light chain complementary determining region of the antibodies. The human immunodeficiency virus HIV antibody is good in specificity, high in biological activity, strong in stability and high in affinity with the human immunodeficiency virus HIV, and can be used for preparing products for detecting the human immunodeficiency virus HIV. The human immunodeficiency virus HIV detection kit provided by the invention has the advantages of being simple and convenient to operate, rapid in reaction, high in sensitivity, strong in specificity, suitable for on-site rapid detection, economical, practical and the like, and can be used for carrying out rapid screening when the virus is epidemic; and a large number of people including suspected cases, asymptomatic infection cases in an incubation period and asymptomatic virus carriers with higher risk can be rapidly, accurately and safely detected in time.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, in particular to human immunodeficiency virus (HIV) antibodies and applications thereof. Background Art

[0002] Acquired immunodeficiency syndrome (AIDS), caused by the human immunodeficiency virus (HIV), has become a major global public health issue, posing a serious threat to public health worldwide. Existing testing and diagnostic technologies are unable to detect acute HIV infection early enough, allowing for the timely implementation of effective interventions. This may be one of the main reasons for the current high HIV prevalence.

[0003] Generally speaking, the clinical symptoms and biological characteristics of HIV infection can be divided into three major phases. The first phase is the acute phase, characterized by rapid viral proliferation and spread in the body, typically occurring between two and four weeks after infection. During this phase, viral replication flares up, and p24 antigen (Ag) levels in the blood are shed and reach a peak. The second phase is the chronic or asymptomatic phase, during which the virus continues to reproduce, but at lower levels. The host immune system also begins to produce antibodies (Abs), and the viral load (VL) decreases to a stable state. As the VL decreases, the p24 antigen level also decreases. This is because the p24 antigen binds to antibodies to form antibody-p24 antigen complexes, which reduce the level of free p24 antigen in the blood. The period from infection to the appearance of antibodies (seroconversion) is called the "window period." If the patient does not receive treatment, as the virus continues to replicate, CD4 cells, which serve as target cells for viral replication, are gradually destroyed, leading to a decline in the CD4 cell count. The third stage is the AIDS stage. As the virus continues to replicate and CD4 cells are consumed, the host immune system is weakened and the level of free p24 antigen in the blood gradually increases.

[0004] Currently, methods for detecting human immunodeficiency virus (HIV) include serological testing, CD4 cell testing, and HIV nucleic acid testing. The most commonly used serological method is antibody testing. However, like CD4 cell testing, antibodies only show significant changes in the second stage, resulting in a long incubation period and limited value for early diagnosis. Nucleic acid testing is currently the most sensitive method for HIV detection, but the high cost of reagents and instrumentation makes it difficult to implement in routine testing. After HIV-1 infection, the earliest detectable viral marker in the blood is the p24 antigen, which can shorten the window for detecting HIV infection to 12-15 days. Therefore, p24 antigen testing is crucial for early diagnosis of HIV infection, prediction of disease progression, prognosis, and screening and evaluation of anti-HIV drug efficacy. Furthermore, combined antigen and antibody testing can predict the stage of infection. Furthermore, self-testing chromatographic methods for detecting p24 antigen in blood offer advantages such as simplicity, low equipment dependency, and short testing time, making them beneficial for early diagnosis. To further shorten the window period, antibodies with strong affinity can be screened for detection to improve the sensitivity of the product. Summary of the Invention

[0005] The objects of the present invention are:

[0006] The first object of the present invention is to provide a method for preparing human immunodeficiency virus (HIV) antibodies, which can prepare antibodies that specifically recognize human immunodeficiency virus (HIV) and achieve efficient antibody pairing.

[0007] The second object of the present invention is to provide human immunodeficiency virus (HIV) antibodies, which can specifically recognize human immunodeficiency virus (HIV).

[0008] The third object of the present invention is to provide the use of the above-mentioned human immunodeficiency virus (HIV) antibody in the preparation of human immunodeficiency virus (HIV) detection products.

[0009] A fourth object of the present invention is to provide a human immunodeficiency virus (HIV) test kit to solve at least one of the above problems.

[0010] In order to achieve the above object, the present invention provides human immunodeficiency virus HIV antibodies, comprising antibody c and / or antibody n;

[0011] The amino acid sequence of the heavy chain complementary determining region of the antibody c includes: CDR1-VH as shown in the amino acid sequence of SEQ ID NO.4, CDR2-VH as shown in the amino acid sequence of SEQ ID NO.5, and CDR3-VH as shown in the amino acid sequence of SEQ ID NO.6;

[0012] The amino acid sequence of the light chain complementary determining region of antibody c includes: CDR1-VL as shown in SEQ ID NO.7, CDR2-VL as shown in SEQ ID NO.8, and CDR3-VL as shown in SEQ ID NO.9;

[0013] The amino acid sequence of the heavy chain complementary determining region of the antibody n includes: CDR1-VH as shown in SEQ ID NO.10, CDR2-VH as shown in SEQ ID NO.11, and CDR3-VH as shown in SEQ ID NO.12;

[0014] The amino acid sequence of the light chain complementary determining region of the antibody n includes: CDR1-VL as shown in SEQ ID NO.13, CDR2-VL as shown in SEQ ID NO.14, and CDR3-VL as shown in SEQ ID NO.15.

[0015] Preferably, the amino acid sequence of the heavy chain variable region of the antibody C is shown in SEQ ID NO.16; the amino acid sequence of the light chain variable region of the antibody C is shown in SEQ ID NO.17;

[0016] Preferably, any of the above items is that the amino acid sequence of the heavy chain variable region of the antibody n is shown as SEQ ID NO.18; the amino acid sequence of the light chain variable region of the antibody n is shown as SEQ ID NO.19.

[0017] Preferably, any of the above items is that the human immunodeficiency virus (HIV) antibody is of mouse origin.

[0018] Preferably, in any of the above items, the light chain of the antibody C has an amino acid sequence as shown in SEQ ID NO.20.

[0019] Preferably, in any of the above items, the heavy chain of the antibody c has an amino acid sequence as shown in SEQ ID NO.21.

[0020] Preferably, in any of the above items, the light chain of the antibody n has an amino acid sequence as shown in SEQ ID NO.22.

[0021] Preferably, in any of the above items, the heavy chain of the antibody n has an amino acid sequence as shown in SEQ ID NO.23.

[0022] Preferably, in any of the above items, the amino acid sequence of the light chain constant region of the antibody C or antibody N is as shown in SEQ ID NO.24.

[0023] Preferably, in any of the above items, the amino acid sequence of the heavy chain constant region of the antibody C or antibody N is as shown in SEQ ID NO.25.

[0024] Preferably, in any of the above items, the nucleotide sequence of the antibody c heavy chain variable region is as shown in SEQ ID NO.26.

[0025] Preferably, in any of the above items, the nucleotide sequence of the antibody C light chain variable region is as shown in SEQ ID NO.27.

[0026] Preferably, in any of the above items, the nucleotide sequence of the heavy chain variable region of the antibody n is as shown in SEQ ID NO.28.

[0027] Preferably, any of the above items is that the nucleotide sequence of the light chain variable region of the antibody n is as shown in SEQ ID NO.29.

[0028] Preferably, in any of the above items, the nucleotide sequence of the light chain constant region of the antibody C or antibody N is as shown in SEQ ID NO.31.

[0029] Preferably, in any of the above items, the nucleotide sequence of the heavy chain constant region of antibody C or antibody N is as shown in SEQ ID NO.30.

[0030] The present invention also provides immunogenic proteins of human immunodeficiency virus (HIV), wherein the immunogenic proteins comprise immunogenic protein C and / or immunogenic protein N.

[0031] Preferably, the immunogenic protein C comprises the amino acid sequence shown in SEQ ID NO.2.

[0032] Preferably, any of the above items is that the immunogenic protein N comprises the amino acid sequence shown in SEQ ID NO.3.

[0033] The present invention also provides the use of the immunogenic protein in preparing any of the above human immunodeficiency virus HIV antibodies.

[0034] Preferably, the antibody C is obtained from the immunogen protein C by immunizing animals.

[0035] Preferably, the antibody n is obtained by immunizing an animal with the immunogen protein N.

[0036] The present invention also provides a method for preparing human immunodeficiency virus HIV antibodies, which comprises two immunogenic proteins;

[0037] The immunogenic protein of human immunodeficiency virus HIV is selected from marker proteins, including immunogenic protein C and immunogenic protein N;

[0038] The immunogen protein C includes the amino acid sequence shown in SEQ ID NO.2; the immunogen protein N includes the amino acid sequence shown in SEQ ID NO.3.

[0039] The present invention also provides a marker protein of human immunodeficiency virus HIV, selected from the p24 protein of the virus, including the amino acid sequence shown in SEQ ID NO.2 and the amino acid sequence shown in SEQ ID NO.3, and the amino acid sequence of the marker protein is shown in SEQ ID NO.1.

[0040] The human immunodeficiency virus HIV antibody provided by the present invention specifically recognizes the marker protein.

[0041] The present invention also provides the use of the human immunodeficiency virus (HIV) antibody in preparing a human immunodeficiency virus (HIV) detection product.

[0042] The present invention also provides a human immunodeficiency virus (HIV) test strip, which comprises a base plate and a sample pad, a marking pad, a detection pad, and a sample suction pad stacked in sequence on the base plate. The marking pad contains any of the above-mentioned human immunodeficiency virus (HIV) antibodies, and the human immunodeficiency virus (HIV) antibodies are marked with a marker.

[0043] Preferably, the marker comprises at least one of colloidal gold, colored microspheres, time-resolved fluorescent microspheres or quantum dot microspheres.

[0044] Preferably, in any of the above items, the particle size of the colloidal gold is 40 to 100 nm; more preferably, it is 40, 50, 60, 80, 100 nm and ranges therebetween.

[0045] Preferably, in any of the above items, the particle size of the colored microspheres is 100 to 300 nm; more preferably, 100, 200, 300 nm and ranges therebetween.

[0046] Preferably, in any of the above items, the particle size of the time-resolved fluorescent microspheres is 100 to 300 nm; more preferably, 100, 200, 300 nm and ranges therebetween.

[0047] Preferably, in any of the above items, the particle size of the quantum dot microspheres is 100 to 300 nm; more preferably, 100, 200, 300 nm and ranges therebetween.

[0048] Preferably, any of the above items has a mass ratio of the human immunodeficiency virus HIV antibody to colloidal gold of (0.04-0.32):1; more preferably, 0.04:1, 0.10:1, 0.15:1, 0.20:1, 0.25:1, 0.30:1, 0.32:1 and ranges therebetween.

[0049] Preferably, any of the above items has a mass ratio of the human immunodeficiency virus HIV antibody to the colored microspheres of (0.1-0.4):1; more preferably, it is 0.1:1, 0.2:1, 0.3:1, 0.4:1 and ranges therebetween.

[0050] Preferably, any of the above items has a mass ratio of the human immunodeficiency virus HIV antibody to the time-resolved fluorescent microspheres of (0.1-0.4):1; more preferably, 0.1:1, 0.2:1, 0.3:1, 0.4:1 and ranges therebetween.

[0051] Preferably, any of the above items has a mass ratio of the human immunodeficiency virus HIV antibody to the quantum dot microspheres of (0.1-0.4):1; more preferably, it is 0.1:1, 0.2:1, 0.3:1, 0.4:1 and ranges therebetween.

[0052] Preferably, any of the above items is provided with a detection line and a quality control line on the detection pad; the detection line is coated with the human immunodeficiency virus HIV antibody; and the quality control line is coated with goat anti-mouse IgG polyclonal antibody.

[0053] Preferably, any of the above items is coated with 0.5-5 mg / mL of the human immunodeficiency virus (HIV) antibody. Further preferably, the test line is coated with 0.5, 1.0, 2.0, 3.0, 4.0, 5.0 mg / mL of the human immunodeficiency virus (HIV) antibody, or a range thereof.

[0054] Preferably, any of the above items is coated with 0.5-5 mg / mL of goat anti-mouse IgG polyclonal antibody. Further preferably, the quality control line is coated with 0.5, 1.0, 2.0, 3.0, 4.0, 5.0 mg / mL of goat anti-mouse IgG polyclonal antibody or a range therebetween.

[0055] Preferably, the diluent for the human immunodeficiency virus (HIV) antibody and the goat anti-mouse IgG polyclonal antibody is a 10-50 mM PB buffer containing trehalose, and each 100 mL of the diluent contains 0.1-1.0 g of trehalose. Further preferably, the diluent is a 10, 20, 30, 40, 50 mM PB buffer containing trehalose, and a range thereof; further preferably, each 100 mL of the diluent contains 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0 g of trehalose, and a range thereof.

[0056] Preferably, any of the above items is provided with a detection line and a quality control line on the detection pad; the detection line is coated with the human immunodeficiency virus HIV antibody; the quality control line is coated with sheep anti-chicken IgY; the labeling pad contains another human immunodeficiency virus HIV antibody, the human immunodeficiency virus HIV antibody on the labeling pad is marked with a marker, and the marker is marked with chicken IgY.

[0057] Preferably, any of the above items is that the human immunodeficiency virus (HIV) antibodies on the labeling pad are different from the human immunodeficiency virus (HIV) antibodies on the detection pad.

[0058] In any of the above, preferably, the HIV antibody on the labeling pad is different from the HIV antibody on the detection pad. Preferably, the HIV antibody on the labeling pad is antibody c, and the HIV antibody on the detection pad is antibody n; preferably, the HIV antibody on the labeling pad is antibody n, and the HIV antibody on the detection pad is antibody c.

[0059] The present invention also provides a human immunodeficiency virus (HIV) test strip and a housing, wherein the human immunodeficiency virus (HIV) test strip is disposed inside the housing.

[0060] Preferably, the housing includes an upper cover and a lower cover that are detachably connected.

[0061] Preferably, any of the above items is that the upper cover is provided with an observation window and a sample addition hole.

[0062] Compared with the prior art, the present invention has the following beneficial effects:

[0063] The present invention provides an antibody preparation method in which the epitopes contained in the immunogen protein are highly conserved and not easily missed, and are dominant epitopes that are easy to generate high-affinity antibodies; and are located at the C-terminus and N-terminus of the p24 antigen, respectively, to facilitate efficient pairing.

[0064] The human immunodeficiency virus (HIV) antibody provided by the present invention has good specificity, high biological activity, strong stability, and high affinity to the human immunodeficiency virus (HIV), and can be used to prepare products for detecting the human immunodeficiency virus (HIV).

[0065] The human immunodeficiency virus (HIV) detection kit provided by the present invention has the advantages of simple operation, rapid reaction, high sensitivity, strong specificity, suitability for rapid on-site detection, and economy and practicality. It is of great significance for the early diagnosis of HIV infection, prediction of disease progression, judgment of prognosis, and screening and evaluation of the efficacy of anti-HIV drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0066] Figure 1 This is the upper cover of the human immunodeficiency virus (HIV) detection kit in preferred embodiment 2 of the present invention.

[0067] Figure 2 This is the lower cover of the human immunodeficiency virus (HIV) detection kit in preferred embodiment 2 of the present invention.

[0068] Figure 3 This is a structural diagram of the human immunodeficiency virus HIV test strip in preferred embodiment 2 of the present invention.

[0069] Figure 4 This is the electrophoresis diagram of the anti-human immunodeficiency virus HIV monoclonal antibody in the preferred embodiment 1 of the present invention.

[0070] Figure 5 This is a display diagram of the detection results in preferred embodiment 3 of the present invention.

[0071] Icons: 1-Observation window; 2-Sample loading hole; 3-Test card loading area; 4-Base plate; 5-Test pad; 6-Sample suction pad; 7-Marking pad; 8-Sample pad; 9-Quality control line; 10-Test line. DETAILED DESCRIPTION

[0072] Below in conjunction with embodiment and example, embodiment of the present invention is described in detail, but those skilled in the art will appreciate that the following embodiment and example are only used to illustrate the present invention, and should not be considered as limiting the scope of the present invention. Based on the embodiment in the present invention, all other embodiments obtained by those of ordinary skill in the art without making creative work premise all fall within the scope of protection of the present invention. Unspecified conditions are carried out according to the conditions of normal conditions or manufacturer's recommendations. Reagents used or instruments not specified by the manufacturer are conventional products that can be purchased commercially.

[0073] It should be noted that the "variable region" or "variable domain" of an antibody refers to the amino-terminal domain of the heavy or light chain of an antibody. The variable domain of the heavy chain can be referred to as "VH". The variable domain of the light chain can be referred to as "VL". These domains are usually the most variable parts of the antibody and contain the antigen binding site. The light or heavy chain variable region is composed of a framework region interrupted by three hypervariable regions called "complementarity determining regions" or "CDRs". The framework region of an antibody, that is, the combined framework region of the constituent light and heavy chains, plays a role in positioning and aligning the CDRs, which are primarily responsible for binding to the antigen.

[0074] "Framework" or "FR" regions refer to the regions of an antibody variable domain excluding those defined as CDRs. Each antibody variable domain framework can be further subdivided into contiguous regions (FR1, FR2, FR3, and FR4) separated by CDRs.

[0075] Typically, the variable regions VL / VH of the heavy and light chains can be obtained by arranging and connecting the following numbered CDRs and FRs in the following combinations: FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4.

[0076] In the present invention, CDR1-VH, CDR2-VH and CDR3-VH refer to the three hypervariable regions of the heavy chain variable region, respectively. Correspondingly, CDR1-VL, CDR2-VL and CDR3-VL refer to the three hypervariable regions of the light chain variable region, respectively.

[0077] Human immunodeficiency virus HIV immunogenic proteins, selected from marker proteins, comprising immunogenic protein C and immunogenic protein N;

[0078] The immunogen protein C includes the amino acid sequence shown in SEQ ID NO.2; the immunogen protein N includes the amino acid sequence shown in SEQ ID NO.3.

[0079] In the first aspect, the present invention provides a marker protein of the human immunodeficiency virus HIV, the amino acid sequence of which is as shown in SEQ ID NO.1; in addition, the present invention provides two immunogenic proteins of the human immunodeficiency virus HIV, comprising immunogenic protein C and immunogenic protein N, the amino acid sequence of which is as shown in SEQ ID NO.2, and the amino acid sequence of which is as shown in SEQ ID NO.3.

[0080] The amino acid sequences shown in SEQ ID NO. 1 to 3 are shown in Table 1

[0081] Table 1

[0082]

[0083] In a second aspect, the present invention provides a human immunodeficiency virus HIV antibody selected from antibody c and / or antibody n;

[0084] The antibody c comprises heavy chain complementary determining regions CDR1-VH, CDR2-VH, and CDR3-VH, whose amino acid sequences are sequentially shown in SEQ ID NOs. 4 to 6, and light chain complementary determining regions CDR1-VL, CDR2-VL, and CDR3-VL, whose amino acid sequences are sequentially shown in SEQ ID NOs. 7 to 9;

[0085] The antibody n includes heavy chain complementary determining regions CDR1-VH, CDR2-VH, and CDR3-VH, whose amino acid sequences are shown in SEQ ID NOs. 10 to 12, and light chain complementary determining regions CDR1-VL, CDR2-VL, and CDR3-VL, whose amino acid sequences are shown in SEQ ID NOs. 13 to 15.

[0086] The amino acid sequences represented by SEQ ID NOs. 4 to 15 are shown in Table 2.

[0087] Table 2

[0088]

[0089]

[0090] The human immunodeficiency virus (HIV) antibody provided by the present invention has good specificity, high biological activity, strong stability, and high affinity to the human immunodeficiency virus (HIV), and can be used to prepare products for detecting the human immunodeficiency virus (HIV).

[0091] In some preferred embodiments, the antibody c comprises a heavy chain variable region and a light chain variable region whose amino acid sequences are sequentially shown in SEQ ID NOs. 16 to 17;

[0092] The antibody n comprises a heavy chain variable region and a light chain variable region whose amino acid sequences are sequentially shown in SEQ ID NOs. 18 to 19;

[0093] Preferably, the human immunodeficiency virus (HIV) antibody is of murine origin.

[0094] The amino acid sequences represented by SEQ ID NOs. 16 to 17 and SEQ ID NOs. 18 to 19 are shown in Table 3.

[0095] Table 3

[0096]

[0097]

[0098] In some preferred embodiments, the light chain of the antibody C has an amino acid sequence as shown in SEQ ID NO. 20. The heavy chain of the antibody C has an amino acid sequence as shown in SEQ ID NO. 21.

[0099] In some preferred embodiments, the light chain of the antibody n has an amino acid sequence as shown in SEQ ID NO. 22. The heavy chain of the antibody n has an amino acid sequence as shown in SEQ ID NO. 23.

[0100] In some preferred embodiments, the amino acid sequence of the light chain constant region of the antibody C or antibody N is shown as SEQ ID NO.24.

[0101] In some preferred embodiments, the amino acid sequence of the heavy chain constant region of the antibody C or antibody N is shown as SEQ ID NO.25.

[0102] The amino acid sequences represented by SEQ ID NO. 20 to SEQ ID NO. 25 are shown in Table 4.

[0103] Table 4

[0104]

[0105]

[0106]

[0107] In a third aspect, the present invention provides the use of the human immunodeficiency virus (HIV) antibody in the preparation of a human immunodeficiency virus (HIV) detection product.

[0108] In a fourth aspect, the present invention provides a human immunodeficiency virus (HIV) test strip, comprising a base plate and a sample pad, a marking pad, a detection pad, and a sample suction pad stacked sequentially on the base plate;

[0109] The marking pad contains the human immunodeficiency virus (HIV) antibody, and the human immunodeficiency virus (HIV) antibody is marked with a marker. The marker can be detected to identify the location or concentration of the marker.

[0110] The human immunodeficiency virus (HIV) test strip provided by the present invention has the advantages of simple operation, rapid reaction, high sensitivity, strong specificity, suitability for on-site rapid detection, and economy and practicality. It is of great significance for the early diagnosis of HIV infection, prediction of disease progression, judgment of prognosis, and screening and evaluation of the efficacy of anti-HIV drugs.

[0111] In some preferred embodiments, the marker includes but is not limited to at least one of colloidal gold, colored microspheres, time-resolved fluorescent microspheres, or quantum dot microspheres;

[0112] Preferably, the particle size of the colloidal gold may be, for example, but not limited to, 40 nm, 60 nm, 80 nm or 100 nm;

[0113] Preferably, the particle size of the colored microspheres, time-resolved fluorescent microspheres and quantum dot microspheres can be, for example, but not limited to, 100 nm, 150 nm, 200 nm, 250 nm or 300 nm;

[0114] Preferably, the mass ratio of human immunodeficiency virus HIV antibody coupled to colloidal gold is: (0.04-0.32):1; the mass ratio of human immunodeficiency virus HIV antibody to colored microspheres, time-resolved fluorescent microspheres or quantum dot microspheres is: (0.1-0.4):1.

[0115] By adjusting the particle size and dosage of the marker, the sensitivity of the test strip can be increased.

[0116] In some preferred embodiments, a detection line (T line) and a quality control line (C line) are provided on the detection pad;

[0117] The test line is coated with 0.5-5 mg / mL of the human immunodeficiency virus HIV antibody;

[0118] The quality control line was coated with 0.5-5 mg / mL goat anti-mouse IgG polyclonal antibody;

[0119] Preferably, the diluent for the human immunodeficiency virus (HIV) antibody and goat anti-mouse IgG polyclonal antibody is a 10-50 mM PB buffer solution containing trehalose, and each 100 mL of the diluent contains 0.1-1.0 g of trehalose.

[0120] In some preferred embodiments, the human immunodeficiency virus (HIV) antibodies on the labeling pad are different from the human immunodeficiency virus (HIV) antibodies on the detection pad.

[0121] For example, the protein on the labeling pad is HIV antibody c, wherein antibody c is human immunodeficiency virus HIV monoclonal antibody 1, and the protein on the detection pad is HIV antibody n, wherein antibody n is human immunodeficiency virus HIV monoclonal antibody 2.

[0122] The test strip provided by the present invention is based on the principle of antigen-antibody reaction, and after labeling human immunodeficiency virus HIV monoclonal antibodies with colloidal gold, colored microspheres, time-resolved fluorescent microspheres, and quantum dots, the marker is solidified on a glass cellulose membrane. Another monoclonal antibody against human immunodeficiency virus HIV is coated on a detection pad (such as an NC membrane), and based on the principle of antigen-antibody reaction, it can be detected by the naked eye or a supporting instrument within the detection time. If human immunodeficiency virus HIV is present in the sample, a double-antibody sandwich structure is formed, forming a band visible to the naked eye or a light intensity signal in the instrument. If human immunodeficiency virus HIV is not present in the sample, no band appears on the NC membrane or there is no light intensity signal in the instrument. Negative or positive judgment is made based on the presence or absence of the signal, or the viral load is pre-judged based on the intensity of the light intensity signal.

[0123] By using the test strip provided by the present invention for testing, the test results can be obtained within 10-30 minutes of the entire process, which is fast and efficient, helping medical personnel to obtain test results in a timely manner, make comprehensive judgments and take timely measures based on the results, avoid panic, and reduce the spread of the epidemic.

[0124] In a fifth aspect, the present invention provides a human immunodeficiency virus (HIV) detection kit, comprising the human immunodeficiency virus (HIV) detection test strip and a shell, wherein the human immunodeficiency virus (HIV) detection test strip is disposed inside the shell.

[0125] The human immunodeficiency virus (HIV) detection kit provided by the present invention contains a human immunodeficiency virus (HIV) detection test strip, and thus has all the beneficial effects of the human immunodeficiency virus (HIV) detection test strip.

[0126] In some preferred embodiments, the housing includes a detachably connected upper cover and a lower cover;

[0127] The upper cover is provided with an observation window and a sample addition hole.

[0128] The shapes of the observation window and the sample addition hole are not specifically limited in the present invention. The observation window can be, for example, square and located above the test line and the quality control line of the test strip for observing the test results. The sample addition hole can be, for example, a circular hole with a diameter of 0.5-1 cm and located above the sample pad.

[0129] Add test sample to the kit:

[0130] It should be noted that, unless otherwise specified in the following examples, the diluent for human immunodeficiency virus (HIV) antibodies and goat anti-mouse IgG polyclonal antibodies is a 20 mM PB buffer solution containing trehalose, with each 100 mL of the diluent containing 0.5 g of trehalose.

[0131] The method for preparing the monoclonal antibody described in the present invention is based on prior art and is only briefly described here. The specific operation method can be carried out according to the operation method described in the prior art:

[0132] (1) Preparation of immunogen:

[0133] A: Based on the amino acid sequence of immunogenic protein C shown in SEQ ID NO. 2 and the amino acid sequence of immunogenic protein N shown in SEQ ID NO. 3 of the present invention, we commissioned GenScript Biotech Co., Ltd. to perform peptide synthesis to obtain immunogenic protein C and immunogenic protein N polypeptides;

[0134] B: KLH-coupled peptides: First, dissolve KLH in coupling buffer, then add immune protein C peptide and immune protein N peptide dropwise to the dissolved KLH buffer, stir overnight, and finally dialyze to obtain KLH-coupled immune protein C peptide and immune protein N peptide for use as immunogens;

[0135] (2) Animal immunization

[0136] A: KLH-coupled immune protein C polypeptide and immune protein N polypeptide were mixed with equal volumes of Freund's adjuvant to an appropriate volume and emulsified completely. Mice were immunized by intraperitoneal injection, with each mouse receiving 50 μg of immunogen in a volume of 100 μL, once a week.

[0137] B: After immunization 4 times, the titer of the antibody in the serum was tested by indirect ELISA method, with the immune protein C polypeptide and immune protein N polypeptide coated respectively. Mice with an OD value greater than 1.0 detected by 16,000-fold serum dilution were screened;

[0138] (3) Preparation of monoclonal antibodies

[0139] A: Splenocytes from screened mice were fused with myeloma cells, and the fused cells were plated and cultured using the limiting dilution method.

[0140] B: Screen the monoclonal cell wells and culture and expand them. The monoclonal cell well with the highest OD value detected in the cell culture supernatant is used as the target hybridoma cell, culture and expand it to obtain the hybridoma cell line and freeze it;

[0141] C: Isolation of antibody variable region genes from hybridoma cells using RT-PCR: After homogenization of the hybridoma cells, add cell lysis buffer for RNA extraction. Precipitate RNA from the aqueous phase with isopropanol. Wash the precipitated RNA after centrifugation to remove impurities, resuspend it, and perform reverse transcription to obtain cDNA.

[0142] D: Mouse-specific primers were used for PCR, and hybridoma cell cDNA was used as a template to amplify the heavy and light chain variable region genes of the antibody. A 50 μL system contained 5 μL cDNA, HotStarTaq Plus enzyme, dNTPs, and 0.5 μM specific primers. PCR amplification was performed under the following conditions: pre-denaturation at 94°C for 5 min; 35 cycles of 94°C for 30 s, 55°C for 30 s, and 72°C for 50 s; and 72°C for 7 min. The obtained PCR products were identified by 1% agarose gel electrophoresis, and the target fragments were recovered and sent for sequencing to obtain the gene fragments of the heavy and light chains of Antibody C and Antibody N. The nucleotide sequence of the heavy chain variable region of Antibody C is shown in SEQ ID NO. 26, the nucleotide sequence of the light chain variable region of Antibody C is shown in SEQ ID NO. 27, the nucleotide sequence of the heavy chain variable region of Antibody N is shown in SEQ ID NO. 28, the nucleotide sequence of the light chain variable region of Antibody N is shown in SEQ ID NO. 29, and the nucleotide sequence of the heavy chain constant region of Antibody C or Antibody N is shown in SEQ ID NO. NO.30, the nucleotide sequence of the light chain constant region of antibody C or antibody N is shown in SEQ ID NO.31, and the antibody variable region amino acid sequences shown in SEQ ID NOs:16-19 are obtained therefrom;

[0143] E: Construction of monoclonal antibody expression vector

[0144] Homologous recombination primers were used to add homologous recombination arms to both ends of the antibody heavy chain variable region gene and the light variable region gene, respectively. The expression plasmid containing the mouse antibody heavy and light chain IgG1 constant regions was linearized using a dual enzyme to generate homologous recombination arms. The variable region gene fragments added with the homologous recombination arms and the linearized plasmid were connected by homologous recombination to form a complete expression vector, the expression vector being pCDNA3.4. The recombinant products were transformed into TOP10 competent Escherichia coli and the plasmids were amplified. The pCDNA3.4-antibody c heavy chain plasmid, pCDNA3.4-antibody c light chain plasmid, pCDNA3.4-antibody n heavy chain plasmid, and pCDNA3.4-antibody n light chain plasmid were obtained, respectively.

[0145] F: Expression and purification of monoclonal antibodies

[0146] The monoclonal antibody heavy and light chain expression plasmids obtained in step E were added to Opti-Mem transfection medium at a ratio of 1:1. After thorough mixing, PEI transfection reagent (4 times the mass of DNA) was added. After mixing, the mixture was placed in the dark at room temperature for 10 minutes, and then added to 293T cells. After incubation for 6 hours, the transfection system was removed, and FreeStyleTM293 expression medium was added. After culturing for 5 days, the cell culture supernatant was collected and the expressed cell culture supernatant was purified by affinity purification (Protein A) to obtain the monoclonal antibody. The specific steps are as follows:

[0147] (1) Centrifuge the expressed antibody supernatant at 2500 × g for 10 min at room temperature to remove the precipitate;

[0148] (2) The affinity purification column containing Protein A was thoroughly washed with 10 volumes of binding buffer;

[0149] (3) The expression supernatant was passed through the purification column at a flow rate of 5 mL / min;

[0150] (4) Wash the purification column thoroughly with 20 times the volume of the purification column binding buffer;

[0151] (5) Elute the purification column with 0.1 M pH = 3.0-3.5 citric acid buffer until the elution peak drops to equilibrium, and adjust the pH to 7.0 with 1 M pH = 9.0 Tris-HCl buffer;

[0152] (6) The purified monoclonal antibody was concentrated using a centrifugal column, PBS was used as the buffer for antibody storage, and the concentration of the concentrated antibody was measured using an ultra-micro UV spectrophotometer.

[0153] In a preferred embodiment of the present invention, the pCDNA3.4-antibody c heavy chain plasmid and the pCDNA3.4-antibody c light chain plasmid were co-transfected at a 1:1 ratio and expressed and purified according to the method in Step F to obtain Antibody C. The pCDNA3.4-antibody n heavy chain plasmid and the pCDNA3.4-antibody n light chain plasmid were expressed and purified according to the method in Step F to obtain Antibody N. The obtained Antibodies C and N were verified by SDS-PAGE and ELISA, respectively, as described in Example 1. Both Antibodies C and N exhibited two characteristic bands of approximately 25 kD and 50 kD, representing the light and heavy chains of IgG, respectively. Indirect ELISA titers confirmed that both Antibodies C and N could specifically recognize HIV.

[0154] The methods used in the present invention for antibody preparation, antibody gene sequencing, construction and identification of monoclonal antibody expression vectors, etc., are all conventional methods in molecular biology. The primers and other related sequences involved can be obtained through existing technologies, such as relevant information recorded in gene databases or existing literature, or can be obtained from the heavy chain target gene fragment or light chain target gene fragment of antibody C or N, such as the nucleotide sequences shown in SEQ ID NO. 26 to SEQ ID NO. 31, and are not described in detail here.

[0155] Preferably, using the above method, the antibody c is prepared from the immunogen protein c; and the antibody n is prepared from the immunogen protein n.

[0156] Example 1

[0157] Human immunodeficiency virus HIV antibody molecular weight and purity verification:

[0158] Human immunodeficiency virus (HIV) antibodies include: human immunodeficiency virus (HIV) monoclonal antibody 1 (antibody c) and human immunodeficiency virus (HIV) monoclonal antibody 2 (antibody n). The variable region sequences of the above antibodies are shown in Table 3.

[0159] A 12% SDS-PAGE gel was prepared according to conventional methods, and 5 μg of the above antibodies were loaded and electrophoresed using a protein molecular weight standard as a reference. The results showed that the two monoclonal antibodies against human immunodeficiency virus HIV showed two characteristic bands of about 25KD and 50KD, which were the light chain and heavy chain of IgG respectively ( Figure 4 ). After scanning and analysis, the antibody content of the strips was above 90%.

[0160] Figure 4 In the table, 1: Anti-1; 2: Anti-2; Anti-1 is anti-human immunodeficiency virus HIV monoclonal antibody 1; Anti-2 is anti-human immunodeficiency virus HIV monoclonal antibody 2.

[0161] Human immunodeficiency virus HIV antibody titer verification:

[0162] The titer of human immunodeficiency virus HIV monoclonal antibody 1 (antibody c) was determined by the indirect ELISA method as follows: the ELISA plate was coated with carrier protein, immune protein C polypeptide, KLH-coupled immune protein C polypeptide, and marker protein, respectively, and the antibody was diluted to 100, 10, 1, and 0.1 ng / mL, respectively, and the affinity of the antibody to different antigens was measured. The results are shown in Table 5.

[0163] Table 5

[0164]

[0165] The titer of human immunodeficiency virus HIV monoclonal antibody 2 (antibody n) was determined by the indirect ELISA method as follows: the ELISA plate was coated with carrier protein, immune protein N polypeptide, KLH-coupled immune protein N polypeptide, and marker protein, respectively, and the antibody was diluted to 100, 10, 1, and 0.1 ng / mL, respectively, and the affinity of the antibody to different antigens was determined. The results are shown in Table 5.

[0166] Table 6

[0167]

[0168] The antibodies used in the following examples are the same as those in Example 1.

[0169] Example 2

[0170] A human immunodeficiency virus HIV detection kit, such as Figures 1 to 3 As shown, the device comprises a human immunodeficiency virus (HIV) test strip and a housing, with the HIV test strip disposed within the housing. The HIV test strip comprises a base plate 4, and a sample pad 8, a marking pad 7, a test pad 5, and a sample suction pad 6 stacked sequentially thereon. The test pad is provided with a test line 10 and a quality control line 9. The housing comprises a detachably connected upper and lower covers. The upper cover is provided with an observation window 1 and a sample loading port 2, while the lower cover is provided with a test card loading area 3.

[0171] Example 3

[0172] Example 3 provides a human immunodeficiency virus (HIV) antigen detection kit prepared by using colloidal gold-labeled antibodies.

[0173] 1 Main Materials

[0174] 1.1 Antibodies: The mouse monoclonal antibodies are antibody C and antibody N described in the present invention, which are labeled and coated respectively; goat anti-mouse IgG antibody: a product of Shenzhen Feipeng Biological Co., Ltd., used for coating the nitrocellulose membrane quality control line.

[0175] 1.2 Nitrocellulose membrane: NC membrane is a product of Sartorius.

[0176] 1.3 Other consumables: PVC boards and other consumables are products of Beacon Labs; commonly used reagents are all analytical grade reagents.

[0177] 1.4 Acquisition of recombinant antigens: Based on the human immunodeficiency virus (HIV) data published by the NCBI database, a protein recombinant antigen with the amino acid sequence shown in SEQ ID NO. 1 was commercially synthesized by GenScript Biotech Co., Ltd.

[0178] 2 Methods

[0179] 2.1 Preparation of colloidal gold labeling pad:

[0180] The steps for preparing the colloidal gold labeling pad are as follows:

[0181] (1) Take 1 mL of colloidal gold solution with a particle size of 40 nm and adjust the pH to 8.5 with 0.2 M K2CO3;

[0182] (2) Add 25 μg of monoclonal antibody 1, with a mass ratio of antibody to colloidal gold particles of 0.1:1. Adjust the rotary shaker to a certain speed, rotate and label at room temperature for 1.5 h, and then add 20 μL of blocking solution;

[0183] (3) Centrifuge at 12000 rpm for 15 min and discard the supernatant;

[0184] (4) Add 100 μL of colloidal gold solution;

[0185] (5) The above concentrate was diluted in a ratio of 1:7, sprayed with gold, and placed in a drying oven at 37°C for 2 hours for later use;

[0186] 2.2NC membrane coating:

[0187] Monoclonal antibody 2 was diluted to 1.5 mg / mL and goat anti-mouse IgG antibody was diluted to 2 mg / mL using 0.02 M PB containing 0.5% trehalose. The test line T and the quality control line C were then drawn on the nitrocellulose membrane using a film sprayer. After coating, the nitrocellulose membrane was dried in an oven at 37°C for 24 h before use.

[0188] 2.3 Assembly of the kit:

[0189] In a drying room, the coated nitrocellulose membrane is placed in the middle of a plastic support plate and pasted. A marker colloidal gold pad is overlapped on one side of the T line of the nitrocellulose membrane (1 / 3 of the colloidal gold pad is overlapped), and a sample pad is overlapped on the other side of the colloidal gold pad (1 / 5 of the colloidal gold pad is overlapped); a sample suction pad is overlapped on one side of the C line of the nitrocellulose membrane (1 / 10 of the sample suction pad is overlapped); and a cutting machine is used to cut the pasted plastic plate into test strips of a certain width, which are then loaded into a test card to form a human immunodeficiency virus HIV antigen detection kit.

[0190] 2.4 Detection:

[0191] Step 1: Take out the test kit and the sample to be tested and equilibrate to room temperature;

[0192] Step 2. Open the sealed aluminum foil bag, take out the test kit and place it flat on the table;

[0193] Step 3: Figure 1 Add 2 drops of sample (about 80-100 μL) to the sample well;

[0194] Step 4: Start the timer and read the result after 10 minutes. Note that if the sample does not undergo lateral chromatography or diafiltration within 1 minute after addition, it may be because the sample is too viscous and needs to be pretreated with saline.

[0195] 3 Results

[0196] Under the action of lateral flow chromatography, when human immunodeficiency virus HIV is present in the sample, the test line will show color, and the quality control line will also show color ( Figure 5 a); When there is no human immunodeficiency virus HIV in the sample, the test line does not show color, and the quality control line shows color ( Figure 5 b); After loading, if the quality control line does not show color, the result will be considered invalid regardless of whether the test line shows color or not ( Figure 5 c in Figure 5 d) in the above.

[0197] 4. Blood samples of HIV patients were tested using the kit provided in Example 3, and the results were consistent with those shown in 3. This indicates the effectiveness of the marker protein, antibody c, and antibody n provided by the present invention in HIV detection.

[0198] Example 4

[0199] Example 4 provides a human immunodeficiency virus (HIV) antigen detection kit prepared by using time-resolved fluorescent microspheres labeled with antibodies.

[0200] A human immunodeficiency virus (HIV) antigen detection kit is prepared using a pair of human immunodeficiency virus (HIV) monoclonal antibodies (antibody C and antibody N) provided by the present invention. The kit comprises a detection card and a test strip. The detection card is divided into an upper cover and a lower base. The test strip has a fluorescent pad embedded with monoclonal antibody 1 (antibody C) labeled with time-resolved fluorescent microspheres, and a detection line coated with monoclonal antibody 2 (antibody N). The double-antibody sandwich method is used to quantitatively detect human immunodeficiency virus (HIV) antigens in a sample.

[0201] 1. Kit preparation process:

[0202] The monoclonal antibody 1 prepared by the method of the present invention is surface labeled with time-resolved fluorescent microspheres. Specific examples are as follows:

[0203] Time-Resolved Fluorescent Microsphere Antibody Labeling: 1 mL of 1% carboxyl time-resolved fluorescent microspheres was added to 9 mL of MES buffer, followed by 25 μL of 10 mg / mL EDC solution and 25 μL of 10 mg / mL NHS solution. The mixture was shaken at room temperature for 30 minutes, and the precipitate was collected by centrifugation. After adding HEPES rehydration solution and ultrasonically dispersing the mixture, 1 mL of 1 mg / mL monoclonal antibody 1 was added. The mixture was shaken at room temperature for 120 minutes, and the precipitate was collected by centrifugation. 1 mL of blocking buffer was then added, and the mixture was shaken at room temperature for 120 minutes. The microsphere precipitate was collected by centrifugation and reconstituted with rehydration solution.

[0204] Preparation of fluorescent pad: After labeling, the time-resolved fluorescent microspheres were diluted with microsphere reconstitution solution and sprayed onto the fluorescent pad using a gold sprayer at a rate of 3 μL / cm and a spraying distance of 6 mm. After spraying, the microspheres were dried at 37°C with low humidity (<30%) for 2 h.

[0205] NC membrane coated CT line: T line uses monoclonal antibody 2 with a concentration of 1.5 mg / mL, and C line uses goat anti-mouse IgG antibody with a concentration of 1 mg / mL. 1 μL / cm line is drawn, and then placed in 37°C low humidity (<30%) for drying for 24 hours.

[0206] Sample pad treatment: The sample pad treatment solution consists of buffer salt, sustained-release agent, cosolvent, blocking agent, etc. The specific formula is 20mM Tris buffer, and each 100mL Tris buffer contains 1g BSA, 0.5g Tween 20, and 2g sucrose. 2 Treat with 1 mL of sample treatment solution. After evenly treating, place in a 37°C low humidity (<30%) oven dry for 2 hours.

[0207] Test strip assembly: Attach the NC film, sample pad, fluorescent pad, and sample pad to the PVC board in that order, with the sample pad and fluorescent pad each pressing 1-2 mm against the NC film, and the sample pad pressing 1-2 mm against the fluorescent pad. After assembly, cut the test strip into widths of 4 ± 0.4 mm and place it in a cartridge. Place the cartridge and desiccant in an aluminum foil bag and seal. Label and box the finished test card.

[0208] Test sample preparation: GenScript Biotech Co., Ltd. commercially synthesized protein recombinant antigen was serially diluted for testing. The results are shown in Table 7.

[0209] Table 7 Recombinant antigen detection results

[0210] Recombinant antigen concentration ng / mL C T t / c Calculate concentration ng / mL 100 15639 62326 3.985 99.38 10 16294 8063 0.495 9.90 1 15329 1057 0.069 0.95 0.1 15209 752 0.049 0.09

[0211] 2 Kit detection process

[0212] Place the test card on a clean, flat surface, draw 80-100 μL of the treated sample and drop it into the sample addition end of the test card. Set up a PBS control group at the same time.

[0213] The standard curve of the test strip was introduced. After 10 minutes, the fluorescent immunoassay analyzer was used to scan the detection area to obtain a fluorescent signal. After the test, the concentration value corresponding to the human immunodeficiency virus HIV antigen was displayed. The test results are shown in Tables 8 and 9 below.

[0214] Table 8 Test results of dilution samples

[0215]

[0216] Table 9 Recombinant antigen test results

[0217]

[0218]

[0219] Example 5

[0220] Example 5 provides a kit for detecting human immunodeficiency virus (HIV) antigens using colored microspheres labeled with antibodies.

[0221] A pair of human immunodeficiency virus (HIV) monoclonal antibodies provided by the present invention is used to prepare a test strip for detecting human immunodeficiency virus (HIV) antigens. The test strip comprises a monoclonal antibody 1 (antibody C) labeled with colored microspheres embedded on a labeling pad, and a detection line coated with a monoclonal antibody 2 (antibody N). The double-antibody sandwich method is used to qualitatively detect human immunodeficiency virus (HIV) antigens in a sample.

[0222] 1. Preparation of test strips:

[0223] 1.1 Colored microspheres labeled with antibodies

[0224] (1) Adjust the pH of 100 nm colored microspheres to 8.0 with 0.1 mol / L K2CO3;

[0225] (2) Monoclonal antibody 1 labeled microspheres: Take 1 mL of the above pH-adjusted solution, add 30 μg of monoclonal antibody 1, the mass ratio of antibody to colored microspheres is 0.3:1, react at room temperature for 1 hour, then centrifuge and discard the supernatant;

[0226] Chicken IgY antibody-labeled microspheres: Add 5 μg of chicken IgY to 1 mL of the pH-adjusted solution (mass ratio of antibody to colored microspheres is 0.05:1). Incubate at room temperature for 1 hour, then centrifuge and discard the supernatant.

[0227] (3) Add 1 mL of 20% BSA to block for 2 h, then centrifuge and discard the supernatant;

[0228] (4) After the above microspheres were redissolved with 100 μL of PH8.0 reconstitution solution, the two microspheres were mixed at a ratio of 5:1, and the mixture was diluted with the reconstitution solution at a ratio of 15%;

[0229] 1.2 Preparation of labeling pad

[0230] The diluted solution of the labeled colored microsphere-antibody complex was sprayed onto the marking pad using a gold sprayer at 7.5 μL / cm and a spraying distance of 6 mm. After spraying, the sample was dried at 37°C with low humidity (<30%) for 2 hours.

[0231] 1.3 Sample pad preparation

[0232] The sample pad treatment solution consists of buffer salt, slow-release agent, cosolvent, blocking agent, etc. The specific formula is 20mM Tris, 1% BSA, 0.5% Tween 20, 2% sucrose. 2 Treat with 1 mL of sample treatment solution. After evenly treating, place in a 37°C low humidity (<30%) oven dry for 2 hours.

[0233] 1.4C / T line coating

[0234] The T line uses monoclonal antibody 2 at a concentration of 1.5 mg / mL, and the C line uses goat anti-chicken IgY antibody at a concentration of 2 mg / mL. 1 μL / cm was used for drawing the line. After the completion, the line was placed in a 37°C low humidity (<30%) oven for 24 h.

[0235] 1.5 Test strip assembly

[0236] Attach the sample pad, NC membrane, and marker pad to the PVC board in this order. Then, press the sample pad and marker pad 1-2 mm below the NC membrane, and press the sample pad 1-2 mm below the marker pad. After assembly, cut the test strips into widths of 4 ± 0.4 mm and place them in the cartridge. Place the cartridge and desiccant in an aluminum foil bag. Label and box the finished test kit.

[0237] 1.6 Test Sample Preparation: GenScript Biotech Co., Ltd. commercially synthesized protein recombinant antigen was serially diluted for testing. The results are shown in Table 10.

[0238] Table 10

[0239] Recombinant antigen concentration ng / mL Interpretation of results 100 Positive 10 Positive 1 Positive 0.1 Negative

[0240] 2. Detection

[0241] Place the test card on a clean, flat surface. Pipette 80-100 μL of the prepared recombinant antigen sample onto the sample tip of the test card. Set up a PBS control group. After 10 minutes, observe the test card window. Two lines indicate positive results, and only one line C indicates negative results. The test results are as follows: Table 11 shows that the antigen match rate for both test strips assembled with monoclonal antibodies was 100%.

[0242] Table 11

[0243] 1× PBS 10 ng / mL Negative Positive Negative Positive Negative Positive

[0244] 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 above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above 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. Human immunodeficiency virus HIV antibody, characterized in that comprising antibody c and / or antibody n; The amino acid sequence of the heavy chain complementary determining region of the antibody c includes: CDR1-VH as shown in the amino acid sequence of SEQ ID NO.4, CDR2-VH as shown in the amino acid sequence of SEQ ID NO.5, and CDR3-VH as shown in the amino acid sequence of SEQ ID NO.6; The amino acid sequence of the light chain complementary determining region of antibody c includes: CDR1-VL as shown in SEQ ID NO.7, CDR2-VL as shown in SEQ ID NO.8, and CDR3-VL as shown in SEQ ID NO.9; The amino acid sequence of the heavy chain complementary determining region of the antibody n includes: CDR1-VH as shown in SEQ ID NO.10, CDR2-VH as shown in SEQ ID NO.11, and CDR3-VH as shown in SEQ ID NO.12; The amino acid sequence of the light chain complementary determining region of the antibody n includes: CDR1-VL as shown in SEQ ID NO.13, CDR2-VL as shown in SEQ ID NO.14, and CDR3-VL as shown in SEQ ID NO.

15.

2. The human immunodeficiency virus (HIV) antibody according to claim 1, wherein The amino acid sequence of the heavy chain variable region of antibody C is shown in SEQ ID NO.16; the amino acid sequence of the light chain variable region of antibody C is shown in SEQ ID NO.17; And / or, the amino acid sequence of the heavy chain variable region of the antibody n is shown as SEQ ID NO.18; the amino acid sequence of the light chain variable region of the antibody n is shown as SEQ ID NO.

19.

3. An immunogenic protein of human immunodeficiency virus (HIV) for use in preparing the human immunodeficiency virus (HIV) antibody according to claim 1 or 2, characterized in that: The immunogenic protein comprises immunogenic protein C and / or immunogenic protein N; the immunogenic protein C comprises the amino acid sequence shown in SEQ ID NO.2; and / or the immunogenic protein N comprises the amino acid sequence shown in SEQ ID NO.

3.

4. A marker protein of human immunodeficiency virus HIV, selected from the p24 protein of the virus, comprising the amino acid sequence shown in SEQ ID NO. 2 and the amino acid sequence shown in SEQ ID NO. 3, characterized in that: The amino acid sequence of the marker protein is shown in SEQ ID NO. 1, and the human immunodeficiency virus HIV antibody according to claim 1 or 2 specifically recognizes the marker protein.

5. Use of the human immunodeficiency virus (HIV) antibody according to claim 1 or 2 in the preparation of a human immunodeficiency virus (HIV) detection product.

6. A human immunodeficiency virus (HIV) test strip, comprising a base plate and a sample pad, a marking pad, a detection pad, and a sample suction pad stacked sequentially on the base plate, characterized in that: The labeling pad contains the human immunodeficiency virus (HIV) antibody according to claim 1 or 2, and the human immunodeficiency virus (HIV) antibody is labeled with a marker.

7. The human immunodeficiency virus (HIV) test strip according to claim 6, wherein: The marker includes at least one of colloidal gold, colored microspheres, time-resolved fluorescent microspheres or quantum dot microspheres.

8. The human immunodeficiency virus (HIV) test strip according to claim 6, wherein: The detection pad is provided with a detection line and a quality control line; the detection line is coated with 0.5-5 mg / mL of the human immunodeficiency virus HIV antibody; The quality control line was coated with 0.5-5 mg / mL goat anti-mouse IgG polyclonal antibody.

9. The human immunodeficiency virus (HIV) test strip according to claim 8, wherein: The human immunodeficiency virus (HIV) antibodies on the labeling pad are different from the human immunodeficiency virus (HIV) antibodies on the detection pad.

10. A human immunodeficiency virus (HIV) detection kit, characterized in that: The invention comprises the human immunodeficiency virus (HIV) detection test strip according to any one of claims 6 to 9 and a shell, wherein the human immunodeficiency virus (HIV) detection test strip is arranged inside the shell.

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