Monoclonal antibody against capsular polysaccharide of klebsiella pneumoniae serotype k64, hybridoma cell line thereof and use thereof

Abstract

A monoclonal antibody against the capsular polysaccharide of Klebsiella pneumoniae serotype K64, a hybridoma cell line thereof and the use thereof. The sequence information of the monoclonal antibody against the capsular polysaccharide of Klebsiella pneumoniae serotype K64 is as set forth in SEQ NO: 1-SEQ NO: 56, and the accession numbers of the hybridoma cell lines are CCTCC: C2023365, CCTCC: C2023366, CCTCC: C2023367, CCTCC: C2024340, CCTCC: C2024341, CCTCC: C2024342 and CCTCC: C2024343, respectively. The provided monoclonal antibody against the capsular polysaccharide of Klebsiella pneumoniae can specifically bind to the capsular polysaccharide of Klebsiella pneumoniae, which is beneficial for the typing of Klebsiella pneumoniae in clinical practice. Both in vivo and in vitro animal experiments have demonstrated that the monoclonal antibody against the capsular polysaccharide of Klebsiella pneumoniae has excellent effects in preventing and treating infections caused by Klebsiella pneumoniae serotype K64, which provides a way to solve the problems of Klebsiella pneumoniae infections and multi-drug resistance, has significant significance in the prevention, diagnosis and treatment of Klebsiella pneumoniae infections, and has an important value for the development of new-generation drugs against Klebsiella pneumoniae.

Description

K64 Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, hybridoma cell line thereof and application thereof Technical Field

[0001] The present application belongs to the field of medical technology, and in particular relates to a monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide of type K64, a hybridoma cell line thereof, and applications thereof. Background Art

[0002] Klebsiella pneumoniae is a Gram-negative (G-) bacterium with an encapsulated bacterium. It primarily colonizes the digestive and respiratory tracts. When the body's immune system is weakened, it can cause lung infections, urinary tract infections, and even bloodstream infections. It is also one of the most common bacterial causes of respiratory tract infections.

[0003] Infections caused by Klebsiella pneumoniae have become a serious global public health problem. Statistics show that Klebsiella pneumoniae infections have an extremely high mortality rate, ranging from 20% to 30% for bloodstream infections and over 50% for bacteremia combined with pneumonia. Treatment of Klebsiella pneumoniae primarily relies on antibiotics. However, with the widespread use of various antimicrobial agents in recent years, Klebsiella pneumoniae that produce extended-spectrum β-lactamases and carbapenemases have become resistant to nearly all available β-lactams, including carbapenems. Over the past decade, the incidence of carbapenem-resistant Klebsiella pneumoniae (CRKP) has increased dramatically worldwide. Capsular polysaccharides are the primary virulence factor of Klebsiella pneumoniae and serve as important protective antigens for the treatment or prevention of Klebsiella pneumoniae infections. Based on the structure of the capsular polysaccharide, Klebsiella pneumoniae can be divided into at least 77 serotypes, also known as K antigens or K types. Different K serotypes exhibit distinct resistance profiles and vary geographically and clinically, potentially leading to different epidemic patterns in nosocomial infections and thus directly impacting clinical treatment options. Among them, K64, despite being less commonly reported, can still cause serious infections, particularly in immunocompromised individuals. Compared with other serotypes, K64 may exhibit unique pathogenicity in certain circumstances. Compared with other serotypes, K64 may have a different antibiotic resistance profile and may exhibit unique pathogenicity in certain environments or populations. However, little research has been conducted on K64 in clinical settings.

[0004] Therefore, there is a need in this field to develop a K64-type Klebsiella pneumoniae capsular polysaccharide monoclonal antibody with high specificity, good affinity and strong binding ability, which is beneficial to the prevention and control of drug-resistant Klebsiella pneumoniae and its clinical application. Summary of the Invention

[0005] To solve the above technical problems, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 1, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 2, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0006] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID: 3, SEQ ID: 4 and SEQ ID: 5, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 3, SEQ ID: 4 and SEQ ID: 5; the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID: 6, SEQ ID: 7 and SEQ ID: 8, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 6, SEQ ID: 7 and SEQ ID: 8.

[0007] In a second aspect, the present application provides a hybridoma cell line, which is named K64-1 and has a deposit number of CCTCC NO: C2023365. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the first aspect.

[0008] In a third aspect, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 9, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 10, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0009] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID: 11, SEQ ID: 12 and SEQ ID: 13, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 11, SEQ ID: 12 and SEQ ID: 13; the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID: 14, SEQ ID: 15 and SEQ ID: 16, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 14, SEQ ID: 15 and SEQ ID: 16.

[0010] In a fourth aspect, the present application provides a hybridoma cell line, which is named K64-23 and has a deposit number of CCTCC NO: C2023366. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the third aspect.

[0011] In a fifth aspect, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 17, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 18, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0012] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID: 19, SEQ ID: 20 and SEQ ID: 21, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 19, SEQ ID: 20 and SEQ ID: 21, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID: 22, SEQ ID: 23 and SEQ ID: 24, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 22, SEQ ID: 23 and SEQ ID: 24.

[0013] In a sixth aspect, the present application provides a hybridoma cell line, which is named K64-24 and has a deposit number of CCTCC NO: C2023367. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the fifth aspect.

[0014] In a seventh aspect, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 25, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 26, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0015] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID: 27, SEQ ID: 28 and SEQ ID: 29, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 27, SEQ ID: 28 and SEQ ID: 29, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID: 30, SEQ ID: 31 and SEQ ID: 32, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 30, SEQ ID: 31 and SEQ ID: 32.

[0016] In an eighth aspect, the present application provides a hybridoma cell line, which is named K64-3 and has a deposit number of CCTCC NO: C2024340. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the seventh aspect.

[0017] In a ninth aspect, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 33 or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 34 or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0018] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID: 35, SEQ ID: 36 and SEQ ID: 37, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 35, SEQ ID: 36 and SEQ ID: 37, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID: 38, SEQ ID: 39 and SEQ ID: 40, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 38, SEQ ID: 39 and SEQ ID: 40.

[0019] In the tenth aspect, the present application provides a hybridoma cell line, the hybridoma cell line is named K64-5, the deposit number of the hybridoma cell line is: CCTCC NO: C2024341, and the hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the ninth aspect.

[0020] In an eleventh aspect, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 41, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 42, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0021] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:43, SEQ ID:44 and SEQ ID:45, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:43, SEQ ID:44 and SEQ ID:45, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:46, SEQ ID:47 and SEQ ID:48, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:46, SEQ ID:47 and SEQ ID:48.

[0022] In the twelfth aspect, the present application provides a hybridoma cell line, which is named K64-8 and has a deposit number of CCTCC NO: C2024342. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the eleventh aspect.

[0023] In a thirteenth aspect, the present application provides a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody, wherein the Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 49 or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 50 or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

[0024] Specifically, the complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID: 51, SEQ ID: 52 and SEQ ID: 53, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 51, SEQ ID: 52 and SEQ ID: 53; the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID: 54, SEQ ID: 55 and SEQ ID: 56, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID: 54, SEQ ID: 55 and SEQ ID: 56.

[0025] In the fourteenth aspect, the present application provides a hybridoma cell line, the hybridoma cell line is named K64-27, the deposit number of the hybridoma cell line is: CCTCC NO: C2024343, and the hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody as described in the fifteenth aspect.

[0026] In a fifteenth aspect, the present application provides a pharmaceutical composition, comprising the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody described in any one of the foregoing, wherein the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody can bind to the Klebsiella pneumoniae capsular polysaccharide target antigen and the non-Klebsiella pneumoniae capsular polysaccharide target antigen, and the pharmaceutical composition further comprises any one or more combinations of pharmaceutically acceptable carriers, excipients or diluents.

[0027] In the sixteenth aspect, the present application provides a kit for detecting K64 type Klebsiella pneumoniae, wherein the kit for detecting K64 type Klebsiella pneumoniae comprises the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody described in any one of the aforementioned methods, and the kit for detecting K64 type Klebsiella pneumoniae further comprises a positive control, a negative control, an antibody diluent, a color developing solution, a stopping solution, a blocking solution or a washing solution, in combination with any one or more of the following.

[0028] In the seventeenth aspect, the present application provides the use of any of the aforementioned Klebsiella pneumoniae capsular polysaccharide monoclonal antibodies in the preparation of K64 type Klebsiella pneumoniae infection disease treatment drugs and / or detection products, wherein the disease comprises one or more symptoms caused by Klebsiella infection, including pneumonia, urinary tract infection, sepsis / bacteremia / sepsis, neonatal sepsis I bacteremia / sepsis, diarrhea, soft tissue infection, post-organ transplant infection, surgical infection, wound infection, lung infection, suppurative liver abscess, lung abscess, cellulitis, necrotizing fasciitis, myositis, endophthalmitis, peritonitis, meningitis, necrotizing meningitis or spondyloarthritis.

[0029] Capsular polysaccharide is an important toxic factor of Klebsiella pneumoniae. This application uses capsular polysaccharide to couple with carrier protein to improve immunogenicity and prepares K64 type Klebsiella pneumoniae capsular polysaccharide-specific monoclonal antibodies using hybridoma technology. The antibodies have good specificity, high affinity, and high binding ability to Klebsiella pneumoniae capsular polysaccharide.

[0030] The beneficial effects of this application are:

[0031] The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody provided in the present application can specifically bind to the capsular polysaccharide of Klebsiella pneumoniae type K64, which is beneficial for the typing of Klebsiella pneumoniae in clinical practice. In vitro and in vivo animal experiments have demonstrated that it has excellent effects in preventing and treating Klebsiella pneumoniae infection, providing a solution to the problems of Klebsiella pneumoniae infection and multidrug resistance, and has significant significance for the prevention, diagnosis and treatment of Klebsiella pneumoniae infection, and is of great value for the development of a new generation of drugs for Klebsiella pneumoniae. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] FIG1 is a graph showing the titer of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody provided by the present application detected by indirect ELISA in Example 2;

[0033] FIG2 is a flow cytometry analysis of the binding of K64 monoclonal antibody, negative control, and K64-type Klebsiella pneumoniae Y8 in Example 4;

[0034] FIG3 is a bactericidal curve of monoclonal antibodies K64-1, K64-23 and K64-24 against a highly virulent strain of Klebsiella pneumoniae (number Y8) in vitro in Example 5;

[0035] FIG4 is a survival curve of the K64-1 antibody preventive trial in Example 6. DETAILED DESCRIPTION

[0036] The following is a clear and complete description of the technical solution of this application. Obviously, the embodiments described are part of the embodiments of this application, not all of them. Based on the embodiments of this application, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of this application.

[0037] The following is the strain deposit information:

[0038] The hybridoma cell line K64-1 (CCTCC NO: C2023365), hybridoma cell line K64-23 (CCTCC NO: C2023366) and hybridoma cell line K64-24 (CCTCC NO: C2023367) provided in this application have all been deposited in the China Center for Type Culture Collection on November 28, 2023.

[0039] The hybridoma cell line K64-3 (CCTCC NO: C2024340), hybridoma cell line K64-5 (CCTCC NO: C2024341), hybridoma cell line K64-8 (CCTCC NO: C2024342) and hybridoma cell line K64-27 (CCTCC NO: C2024343) provided in this application have all been deposited in the China Center for Type Culture Collection on September 25, 2024.

[0040] Deposit address: Wuhan University, Wuhan, Hubei Province.

[0041] Deposit name and deposit number:

[0042] Hybridoma cell line K64-1: Hybridoma cell line K64-1, CCTCC NO: C2023365.

[0043] Hybridoma cell line K64-23: Hybridoma cell line K64-23, CCTCC NO: C2023366.

[0044] Hybridoma cell line K64-24: Hybridoma cell line K64-24, CCTCC NO: C2023367.

[0045] Hybridoma cell line K64-3: Hybridoma cell line K64-3. CCTCC NO: C2024340.

[0046] Hybridoma cell line K64-5: Hybridoma cell line K64-5. CCTCC NO: C2024341.

[0047] Hybridoma cell line K64-8: Hybridoma cell line K64-8, CCTCC NO: C2024342.

[0048] Hybridoma cell line K64-27: Hybridoma cell line K64-27, CCTCC NO: C2024343.

[0049] In Examples 4 and 5 of the present application, the light chain variable region (VL) sequence of the negative control antibody used (as shown in SEQ ID NO: 57) is:

[0050]

[0051] The negative control antibody heavy chain variable region (VH) sequence used (as shown in SEQ ID NO: 58):

[0052]

[0053] Example 1: Preparation of monoclonal antibodies against Klebsiella pneumoniae K64 capsular polysaccharide

[0054] 1) Establishment of a cell line producing monoclonal antibodies against Klebsiella pneumoniae K64 capsular polysaccharide

[0055] The K64 capsular polysaccharide-protein conjugate was obtained by chemically coupling the capsular polysaccharide extracted from the fermentation of Klebsiella pneumoniae K64 (GDMCC No: 62132) with a carrier protein (diphtheria toxin-free variant CRM197) using commonly used polysaccharide activation methods, such as the cyanogen bromide method (reference US6375846B1), 1-cyano-4-dimethylamino arsenic tetrafluoroborate (CDAP) (EP0720485), and the periodate oxidation method (US4711779). The K64 polysaccharide protein conjugate was added to Freund's adjuvant and immunized into SPF-grade BALB / c mice multiple times. When the immune titer was greater than 1:300,000, the spleens were harvested and hybridoma technology was used. After PEG1500 fusion, indirect ELISA screening of positive cell supernatants, and multiple subcloning, three monoclonal cell lines were obtained: hybridoma cell line K64-1 (CCTCC NO: C2023365), hybridoma cell line K64-23 (CCTCC NO: C2023366), hybridoma cell line K64-24 (CCTCC NO: C2023367), hybridoma cell line K64-3 (CCTCC NO: C2024340), hybridoma cell line K64-5 (CCTCC NO: C2024341), hybridoma cell line K64-8 (CCTCC NO: C2024342), and hybridoma cell line K64-27 (CCTCC NO: C2024343). NO: C2024343), and its pure strain was verified by sequencing. 。

[0056] After sequencing, the sequence information of 7 Klebsiella pneumoniae capsular polysaccharide monoclonal antibodies are as follows (SEQ NO: 1 to SEQ NO: 56):

[0057] K64-1 light chain variable region (VL) sequence (starting from FR-L1):

[0058]

[0059] K64-1 heavy chain variable region (VH) sequence (starting from FR-H1):

[0060]

[0061] Complementarity determining region 1 (LCDR1) sequence of K64-1 light chain:

[0062]

[0063] Complementarity determining region 2 (LCDR2) sequence of K64-1 light chain:

[0064]

[0065] Complementarity determining region 3 (LCDR3) sequence of K64-1 light chain:

[0066]

[0067] Complementarity determining region 1 (HCDR1) sequence of K64-1 heavy chain:

[0068]

[0069] Complementarity determining region 2 (HCDR2) sequence of K64-1 heavy chain:

[0070]

[0071] Complementarity determining region 3 (HCDR3) sequence of K64-1 heavy chain:

[0072]

[0073] K64-23 light chain variable region (VL) sequence (starting from FR-L1):

[0074]

[0075] K64-23 heavy chain variable region (VH) sequence (starting from FR-H1):

[0076]

[0077] Complementarity determining region 1 (LCDR1) sequence of the K64-23 light chain:

[0078]

[0079] Complementarity determining region 2 (LCDR2) sequence of the K64-23 light chain:

[0080]

[0081] Complementarity determining region 3 (LCDR3) sequence of the K64-23 light chain:

[0082]

[0083] Complementarity determining region 1 (HCDR1) sequence of K64-23 heavy chain:

[0084]

[0085] Complementarity determining region 2 (HCDR2) sequence of K64-23 heavy chain:

[0086]

[0087] Complementarity determining region 3 (HCDR3) sequence of K64-23 heavy chain:

[0088]

[0089] K64-24 light chain variable region (VL) sequence (starting from FR-L1):

[0090]

[0091] K64-24 heavy chain variable region (VH) sequence (starting from FR-H1):

[0092]

[0093] Complementarity determining region 1 (LCDR1) sequence of the K64-24 light chain:

[0094]

[0095] Complementarity determining region 2 (LCDR2) sequence of the K64-24 light chain:

[0096]

[0097] Complementarity determining region 3 (LCDR3) sequence of K64-24 light chain:

[0098]

[0099] Complementarity determining region 1 (HCDR1) sequence of K64-24 heavy chain:

[0100]

[0101] Complementarity determining region 2 (HCDR2) sequence of K64-24 heavy chain:

[0102]

[0103] Complementarity determining region 3 (HCDR3) sequence of K64-24 heavy chain:

[0104]

[0105] K64-3 variable chain (VL) sequence (starting from FR-L1):

[0106]

[0107] K64-3 heavy chain variable region (VH) sequence (starting from FR-H1):

[0108]

[0109] Complementarity determining region 1 (LCDR1) sequence of K64-3 light chain:

[0110]

[0111] Complementarity determining region 2 (LCDR2) sequence of K64-3 hind linker:

[0112]

[0113] Complementarity determining region 3 (LCDR3) sequence of K64-3 light chain:

[0114]

[0115] Complementarity determining region 1 (HCDR1) sequence of K64-3 heavy chain:

[0116]

[0117] Complementarity determining region 2 (HCDR2) sequence of K64-3 heavy chain:

[0118]

[0119] Complementarity determining region 3 (HCDR3) sequence of K64-3 heavy chain:

[0120]

[0121] K64-5 light chain variable region (VL) sequence (starting from FR-L1):

[0122]

[0123] K64-5 heavy chain variable region (VH) sequence (starting from FR-H1):

[0124]

[0125] Complementarity determining region 1 (LCDR1) sequence of K64-5 light chain:

[0126]

[0127] Complementarity determining region 2 (LCDR2) sequence of the K64-5 chain:

[0128]

[0129] Complementarity determining region 3 (LCDR3) sequence of K64-5 light chain:

[0130]

[0131] Complementarity determining region 1 (HCDR1) sequence of K64-5 heavy chain:

[0132]

[0133] Complementarity determining region 2 (HCDR2) sequence of K64-5 heavy chain:

[0134]

[0135] Complementarity determining region 3 (HCDR3) sequence of K64-5 heavy chain:

[0136]

[0137] K64-8 variable chain (VL) sequence (starting from FR-L1):

[0138]

[0139] K64-8 heavy chain variable region (VH) sequence (starting from FR-H1):

[0140]

[0141] Complementarity determining region 1 (LCDR1) sequence of K64-8 light chain:

[0142]

[0143] Complementarity determining region 2 (LCDR2) sequence of K64-8 light chain:

[0144]

[0145] Complementarity determining region 3 (LCDR3) sequence of K64-8 light chain:

[0146]

[0147] Complementarity determining region 1 (HCDR1) sequence of K64-8 heavy chain:

[0148]

[0149] Complementarity determining region 2 (HCDR2) sequence of K64-8 heavy chain:

[0150]

[0151] Complementarity determining region 3 (HCDR3) sequence of K64-8 heavy chain:

[0152]

[0153] K64-27 light chain variable region (VL) sequence (starting from FR-L1):

[0154]

[0155] K64-27 heavy chain variable region (VH) sequence (starting from FR-H1):

[0156]

[0157] Complementarity determining region 1 (LCDR1) sequence of the K64-27 light chain:

[0158]

[0159] Complementarity determining region 2 (LCDR2) sequence of the K64-27 light chain:

[0160]

[0161] Complementarity determining region 3 (LCDR3) sequence of the K64-27 light chain:

[0162]

[0163] Complementarity determining region 1 (HCDR1) sequence of K64-27 heavy chain:

[0164]

[0165] Complementarity determining region 2 (HCDR2) sequence of K64-27 heavy chain:

[0166]

[0167] Complementarity determining region 3 (HCDR3) sequence of K64-27 heavy chain:

[0168]

[0169] As used herein, the term "CDR" or "complementarity determining region" refers to the non-contiguous antigen binding sites found within the variable regions of heavy and light chain polypeptides. In this application, the amino acid sequences of the CDRs listed above are all shown according to the Kabat definition (the claims of the present invention also show sequences according to the Kabat definition). However, it is well known in the art that antibody CDRs can be defined in the art using a variety of methods. In the literature, Kabat et al., J. Biol. Chem. 252: 6609-6616 (1977); Kabat et al., USDept. of Health and Human Services, "Sequences of proteins of immunological interest" (1991); Chothia et al, J. Mol. Biol. 196: 901-917 (1987); A1-Lazikani B. et al. al., J.Mal.Biol., 273: 927-948 (1997); MacCallum et al, J.Mal.Biol. 262: 732-745 (1996); Abhinandan and Martin, Mal.Immunol., 45: 3832-3839 (2008); Lefranc MPet These specific regions have been described in, for example, Desmond et al., Dev. Comp. J. Am. Biol., 27:55-77 (2003); and Honegger and Plitikthun, J. Mal. Biol., 309:657-670 (2001), wherein these definitions include overlap or subsets of amino acid residues when compared to one another. However, any of the definitions used to designate a CDR of an antibody or grafted antibody or variant thereof is included within the scope of the term as defined and used herein. Algorithms and binding interfaces for CDR prediction are known in the art, such as Abhinandan and Martin, Mal. Immunol., 45: 3832-3839 (2008); Ehrenmann F. et al, Nucleic Acids Res.. 38: D301-D307 (2010); and Adolf-Bryfogle J. et al., Nucleic Acids Res., 43: D432-D438 (2015). The contents of the references cited in this paragraph are incorporated herein by reference in their entirety for use in this application and in one or more claims that may be included in this application.

[0170] As used herein, "monoclonal antibody" refers to a group of homologous antibodies that participate in the highly specific recognition and binding of a single antigenic determinant or epitope, covering complete and full-length monoclonal antibodies as well as antibody fragments (such as Fab, Fab', F(ab')2, Fv), single-chain (scFv) mutants, fusion proteins comprising antibody portions, and any other modified immunoglobulin molecules comprising antigen recognition sites. In addition, "monoclonal antibody" refers to such antibodies prepared in any number of ways (including but not limited to by hybridomas, phage selection, recombinant expression, and transgenic animals). In humans, this category includes IgG1, IgG2, IgG3, and IgG4. In mice, this category includes IgG1, IgG2a, IgG2b, and IgG3. It is known in the art that the antigen-binding function of an antibody can be performed by fragments of a full-length antibody. The monoclonal antibodies or antigen-binding fragments described in the claims of the present invention include Fc fragments in the above-mentioned categories of antibodies.

[0171] 2) Preparation and purification of monoclonal antibodies against Klebsiella pneumoniae K64 capsular polysaccharide in ascites

[0172] The monoclonal antibody cells were expanded and cultured, and 0.5 mL of the monoclonal antibody with a concentration of 1×10 6 100 cells / mL were added to BALB / c mice sensitized with paraffin oil 7 days in advance. After 7-10 days, the mice were observed to have a noticeable bulge in the abdomen, and the ascites was collected for purification.

[0173] The mouse ascites was diluted with PBS, and 50% final concentration of ammonium sulfate was added to precipitate the crude product. The monoclonal antibody with a purity greater than 95% was obtained after Protein A affinity column chromatography and ion exchange chromatography. The monoclonal antibody was sterile filtered through a 0.22 μm filter membrane and stored in PBS at pH 7.0-7.4.

[0174] Example 2: Titer Level Detection of Monoclonal Antibodies to Klebsiella Pneumoniae K64 Capsular Polysaccharide

[0175] After cell fusion, screening, and several subcloning steps, seven monoclonal antibody cell lines were obtained. The results of ELISA tests for binding of their cell culture supernatants to K64-type capsular polysaccharide are shown in Table 1 below.

[0176] Table 1

[0177]

[0178] As shown in FIG1 , the titer of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody provided in the present application was detected by indirect ELISA method, and it can be seen that the supernatants of the seven cell strains all had good binding affinity with the K64 capsular polysaccharide.

[0179] Example 3: ELISA detection of binding specificity of monoclonal antibodies to Klebsiella pneumoniae K64 capsular polysaccharide

[0180] The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody provided in this application was analyzed by indirect ELISA to react with K47, K64, K19, K1, K2, K38 and K57 capsular polysaccharides to evaluate its specificity. Klebsiella pneumoniae capsular polysaccharides of different serotypes were coated on an ELISA plate at 37°C for 3 hours. After blocking with 1% BSA for 1 hour, an appropriate amount of K64 Klebsiella pneumoniae capsular polysaccharide monoclonal antibody was added to react with the coated polysaccharide, 50 μL per well, and incubated overnight. After washing the plate, AP-labeled goat anti-mouse secondary antibody was diluted 1:30,000 and added to the ELISA plate, 100 μL per well, incubated for 2 hours, and then washed. Then, 1 mg / mL PNPP-Na colorimetric substrate was added, 100 μL per well, and after incubation for 2 hours, 3M NaOH was added to 50 μL wells to stop the reaction, and the OD405 value was read on the machine.

[0181] The results of the binding of the 7 K64 Klebsiella pneumoniae capsular polysaccharide monoclonal antibodies provided in this application to Klebsiella capsular polysaccharides of different serotypes are shown in Table 2 below:

[0182] Table 2

[0183]

[0184] It can be seen from Table 2 above that the seven K64 Klebsiella pneumoniae capsular polysaccharide monoclonal antibodies provided in the present application are specific to K64 capsular polysaccharide and have no cross-reaction with other serotypes of Klebsiella capsular polysaccharides.

[0185] Example 4: Flow Cytometry Detection of Binding of Klebsiella pneumoniae K64 Capsular Polysaccharide Monoclonal Antibodies to Klebsiella pneumoniae K64

[0186] K64-serotype highly virulent Klebsiella pneumoniae Y8 (Li, J.; Sheng, Y.; Ma, R.; Xu, M.; Liu, F.; Qin, R.; Zhu, M.; Zhu, X.; He, P. Identification of a Depolymerase Specific for K64-Serotype Klebsiella pneumoniae: Potential Applications in Capsular Typing and Treatment. Antibiotics 2021, 10, 144.) was inoculated into 5 nL of LB medium and cultured at 37°C and 250 rpm to an OD value of 0.7. 0.2 mL of the bacterial solution was centrifuged at 8000 g for 10 min, and the precipitate was collected, washed twice with PBS, and resuspended in 0.2 mL of LB medium. PBS; then fixed with 4% paraformaldehyde for 10 minutes, blocked with 2% BSA for 1 hour after washing, and then incubated with K64 capsular polysaccharide monoclonal antibody overnight. After washing, anti-mouse fluorescent antibody (brand-CST, product number-#4410S) was added and incubated in the dark for 1 hour. After washing with PBS, the cells were resuspended in PBS and detected by flow cytometry.

[0187] After culturing the highly virulent Klebsiella pneumoniae strain Y8, the binding ability was analyzed by flow cytometry using K64-type monoclonal antibodies. The test results are shown in Figure 2. Seven K64-type monoclonal antibodies were tested for binding to the K64-type highly virulent Klebsiella pneumoniae strain Y8. The results showed that all seven K64-type monoclonal antibodies could bind to Y8, while the negative control did not bind to Y8, indicating that these seven K64-type capsular polysaccharide monoclonal antibodies can be used in the diagnosis and typing of clinical strains.

[0188] Example 5: Bactericidal test of monoclonal antibodies against Klebsiella pneumoniae K64 capsular polysaccharide

[0189] This example shows the K64 type Klebsiella pneumoniae capsular polysaccharide monoclonal antibody opsonization swallowing bactericidal test provided by the present application, specifically: K64 type highly virulent Klebsiella pneumoniae Y8 strain was diluted 10 5 CFU / ml, and added to a 96-well cell working plate at 10 μL / well. K64 type capsular polysaccharide monoclonal antibody was serially diluted and added to the above cell working plate at 20 μL / well. The bacteria and antibody were incubated at 700 rpm / min for 30 minutes. The HL-60 cells differentiated by DMF were washed with HBSS buffer and adjusted to a concentration of 1×10 71×107 cells / ml was then mixed with diluted complement at a volume ratio of 1:4. 50 μL of this mixture was added to a 96-well plate. The 96-well plate was placed on a mixer and placed in a 5% CO2 incubator at 37°C with shaking for 45 minutes. After opsonophagocytosis was stopped, the plate was spotted onto a blood agar plate and incubated overnight in a CO2 incubator. The bactericidal efficiency of each antibody dilution was calculated.

[0190] As shown in Figure 3, K64-type capsular polysaccharide monoclonal antibodies K64-1, K64-23, and K64-24 had good bactericidal effects on the highly virulent strain Y8 in vitro, with the highest bactericidal rates reaching 83.3% and 86.7%, respectively, which were significantly different from the negative control antibodies.

[0191] Example 6: Prophylactic Test of Monoclonal Antibodies to Klebsiella Pneumoniae K64 Capsular Polysaccharide

[0192] This example shows the protective test of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody provided by the present application. Specifically, the K64-1 Klebsiella pneumoniae capsular polysaccharide monoclonal antibody was administered by intraperitoneal injection to 6 mice per group, with each mouse receiving 0.025 mg. PBS was used as a negative control. Four hours later, the highly virulent Klebsiella pneumoniae strain Y8 was injected at a concentration of 7.38×10 6 Balb / c mice (n=6) were infected intraperitoneally with the bacterial count of CFU. The survival rate of the mice was observed 10 days after infection, and a survival curve was drawn.

[0193] As shown in Figure 4, it can be seen that the amount of Klebsiella pneumoniae type 64 (strain number Y8) was 7.38×10 6 After CFU challenge, the survival rate of the control group was 0%, while that of the monoclonal antibody group was 100%, with statistical difference (p<0.05).

[0194] In summary, the above embodiments are merely preferred embodiments of the present application and are not intended to limit the scope of protection of the present application. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principles of the present application should be included in the scope of protection of the present application.

Claims

1. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 1, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 2, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

2. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 1, characterized in that The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:3, SEQ ID:4 and SEQ ID:5, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:3, SEQ ID:4 and SEQ ID:5; the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:6, SEQ ID:7 and SEQ ID:8, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:6, SEQ ID:7 and SEQ ID:

8.

3. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-1, and the deposit number of the hybridoma cell line is CCTCC NO: C2023365. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 1.

4. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 9, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 10, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

5. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 4, characterized in that The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:11, SEQ ID:12 and SEQ ID:13, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:11, SEQ ID:12 and SEQ ID:13; the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:14, SEQ ID:15 and SEQ ID:16, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:14, SEQ ID:15 and SEQ ID:

16.

6. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-23, and the deposit number of the hybridoma cell line is CCTCC NO: C2023366. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 4.

7. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 17, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 18, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

8. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 7, characterized in that: The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:19, SEQ ID:20 and SEQ ID:21, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:19, SEQ ID:20 and SEQ ID:21, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:22, SEQ ID:23 and SEQ ID:24, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:22, SEQ ID:23 and SEQ ID:

24.

9. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-24, and the deposit number of the hybridoma cell line is CCTCC NO: C2023367. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 7.

10. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 25, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 26, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

11. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 10, characterized in that: The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:27, SEQ ID:28 and SEQ ID:29, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:27, SEQ ID:28 and SEQ ID:29, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:30, SEQ ID:31 and SEQ ID:32, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:30, SEQ ID:31 and SEQ ID:

32.

12. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-3, and the deposit number of the hybridoma cell line is CCTCC NO: C2024340. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 10.

13. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO: 33, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO: 34, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

14. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 13, characterized in that The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:35, SEQ ID:36 and SEQ ID:37, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:35, SEQ ID:36 and SEQ ID:37, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:38, SEQ ID:39 and SEQ ID:40, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:38, SEQ ID:39 and SEQ ID:

40.

15. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-5, and the deposit number of the hybridoma cell line is CCTCC NO: C2024341. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 13.

16. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO:41, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO:42, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

17. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 16, characterized in that The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:43, SEQ ID:44 and SEQ ID:45, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:43, SEQ ID:44 and SEQ ID:45, and the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:46, SEQ ID:47 and SEQ ID:48, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:46, SEQ ID:47 and SEQ ID:

48.

18. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-8, and the deposit number of the hybridoma cell line is CCTCC NO: C2024342. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 16.

19. A monoclonal antibody against Klebsiella pneumoniae capsular polysaccharide, characterized in that: The Klebsiella pneumoniae is K64 type Klebsiella pneumoniae, and the sequence of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody comprises a light chain variable region and a heavy chain variable region, the amino acid sequence of the light chain variable region is as shown in SEQ NO:49, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto, and the amino acid sequence of the heavy chain variable region is as shown in SEQ NO:50, or a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity thereto.

20. The Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 19, characterized in that The complementarity determining regions of the light chain variable region comprise three sequences as shown in SEQ ID:51, SEQ ID:52 and SEQ ID:53, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:51, SEQ ID:52 and SEQ ID:53; the complementarity determining regions of the heavy chain variable region comprise three sequences as shown in SEQ ID:54, SEQ ID:55 and SEQ ID:56, or sequences having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity compared to any one of SEQ ID:54, SEQ ID:55 and SEQ ID:

56.

21. A hybridoma cell line, characterized in that The hybridoma cell line is named K64-27, and the deposit number of the hybridoma cell line is CCTCC NO: C2024343. The hybridoma cell line is used to prepare the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to claim 19.

22. A pharmaceutical composition, characterized in that The pharmaceutical composition comprises the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to any one of claims 1, 4, 7, 10, 13, 16 and 19, and further comprises any one or more combinations of pharmaceutically acceptable carriers, excipients or diluents.

23. A kit for detecting Klebsiella pneumoniae K64, characterized in that: The kit for detecting Klebsiella pneumoniae type K64 comprises a Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to any one of claims 1, 4, 7, 10, 13, 16 and 19, and the kit for detecting Klebsiella pneumoniae type K64 further comprises a positive control, a negative control, an antibody diluent, a color developing solution, a stop solution, a blocking solution or a washing solution, any one or more combinations thereof.

24. Use of the Klebsiella pneumoniae capsular polysaccharide monoclonal antibody according to any one of claims 1, 4, 7, 10, 13, 16 and 19 in the preparation of a drug for treating K64 type Klebsiella pneumoniae infection and / or a detection product.

Images