25 results about "Splenocyte" patented technology

The invention discloses preparation of a human CD56 immune phage display antibody library, which comprises the following steps: S1, animal immunization: expressing recombinant hCD56 protein in escherichia coli, then immunizing a Balb / c mouse, and detecting immune titer; s2, splenocyte preparation: extracting RNA (Ribonucleic Acid) of immune mouse spleen cells, and performing reverse transcription to obtain cDNA (Complementary Deoxyribonucleic Acid); s3, preparation of an initial bacteriophage display library: amplifying a heavy chain variable region and a light chain variable region by using a PCR technology, and then assembling to form scFv; constructing scFv phagocytids, electrically transforming the scFv phagocytids into TG1 competent cells, and constructing a bacterial library; s4, constructing a phage single-chain library of the hCD56: infecting the bacterial library by using a helper phage M13K07, and constructing a phage single-chain antibody library; and S5, elutriation and screening of a phage library: specifically enriching a phage single-chain antibody library, screening positive clones from the phage single-chain antibody library, and determining a sequence. According to the invention, the human hCD56 antibody scFv sequence fragment can be screened out, and the research and development period is shortened.

The invention discloses a monoclonal antibody targeting all CD45 subtypes and application of the monoclonal antibody, and belongs to the field of monoclonal antibody preparation. The hybridoma cell strain capable of stably secreting all anti-human CD45 subtype antibodies is successfully obtained by stably expressing short and long molecular subtypes of human CD45 extracellular regions in L929 mouse fibroblasts, immunizing BALB / c mice with L929 cells expressing the two molecular subtypes of human CD45, and fusing splenocytes with SP20 myeloma cells after the serum titer reaches the standard. And the monoclonal antibody targeting all subtypes of CD45 is obtained. The monoclonal antibody provided by the invention shows a high-affinity binding characteristic with all human CD45 molecular subtypes, has important application value in the aspect of human CD45 molecular detection, and can be used as a therapeutic antibody in the fields of tumor immunotherapy, autoimmune disease treatment, transplant rejection resistance and the like.

The present invention relates to a composition for enhancing immunity, containing, as an active ingredient, a polysaccharide enzyme-treated product derived from a mixed extract of Angelica gigas, Cnidium officinale, and Paeonia lactiflora, or a novel purified polysaccharide fraction thereof. The composition for enhancing immunity according to the present invention, comprising the polysaccharide enzyme-treated product of the mixed extract of Angelica gigas, Cnidium officinale, and Paeonia lactiflora or the novel purified polysaccharide fraction thereof, is composed of specific types and contents of sugars and significantly increases the activity of macrophages, splenocytes, and NK cells upon application thereto, thereby being applicable as an immunopotentiator or a composition for enhancing immunity.

Described herein is an engineered viral subunit vaccine encoding ASFV capsid proteins (e.g., P72 and Penton) for use in pigs. The vaccine with either P72 or Penton tested in mice elicited significantly higher levels of antigen-specific IgM and IgG, increased frequency of antibody-secreting cells in the bone marrow, and higher quality germinal centers in the spleens of immunized mice. Enhanced T cell responses were also observed, with higher levels of IFN-γ and TNF-α in splenocytes. Immunogenicity assessments in pigs further supported these findings, with both MB-P72 and MB-PN180Q inducing robust B cell and T cell responses. These findings show that expression of capsid proteins P72 and Penton in membrane-bound forms via mRNA preserves their native multimeric structure and enhances their immunogenicity and provides a basis for an effective ASFV vaccine.

To conduct detection of Salmonella, serotype transversely and specifically relative to other strains.SOLUTION: A heat-killed bacterial cell of Salmonella and antigen liquid containing an adjuvant are inoculated in a BALB / c mouse. After the inoculation, splenocytes or lymph node cells of the mouse and P3U1 are fused to prepare a hybridoma, and therefrom, a hybridoma for producing a monoclonal IgG antibody showing Salmonella specificity is selected, whereby three hybridomas for producing superior Salmonella specific antibodies are successfully acquired. Next, these monoclonal antibodies are used to detect Salmonella of a plurality serotypes and other strains according to sandwich ELISA, thereby obtaining a result in which Salmonella is detected serotype transversely, whereas other strains are not detected.SELECTED DRAWING: Figure 1

The invention belongs to the technical field of bioengineering, and relates to a construction method and application of a multiple myeloma extramedullary lesion mouse model. The method comprises the following steps: performing intravenous injection on a Vk12653 tumor cell suspension at the tail of a C57BL / 6J mouse to construct a Vk * MYC myeloma mouse; feeding Vk * MYC myeloma mice to obtain spleen cells infiltrated by tumors; a fibrinogen alpha chain is knocked out from the whole body of a C57BL / 6J mouse, and splenocytes are transplanted; and continuously culturing to obtain the multiple myeloma extramedullary lesion mouse model. The transplanted animal model provided by the invention can be used for effectively screening multiple myeloma extramedullary metastasis drugs and testing multiple drugs and drug combination, meanwhile, the in-vivo mechanism of multiple myeloma extramedullary metastasis can be more comprehensively and deeply researched by using the model, and a basic support is provided for developing new specific drugs.

The invention provides a PEDV (porcine epidemic diarrhea virus) N protein monoclonal antibody as well as a preparation method and application thereof, and the method comprises the following steps: performing prokaryotic expression on PEDV N protein, and purifying to obtain the PEDV N protein; emulsifying the purified PEDV N protein and an isopyknic Freund's adjuvant, injecting the emulsified PEDV N protein into 8-week-old female BALB / c mice at multiple points, and immunizing once every two weeks; the method comprises the following steps: uniformly mixing splenocytes of a mouse subjected to boosted immunization for 3 days with SP2 / 0 cells in a logarithmic phase according to a ratio of 5: 1, carrying out cell fusion under the action of PEG 1500, and culturing a fused cell mixed solution by using an HAT culture medium; screening out positive pores with strong antibody secretion capacity by adopting an indirect ELISA (Enzyme-Linked Immunosorbent Assay) method, and carrying out subcloning by adopting a limited dilution method to obtain six cell strains for secreting antibodies; the PEDV N protein mAb is prepared by adopting an in-vivo ascites inducing method. The prepared PEDV N protein monoclonal antibody can be specifically and efficiently combined, and can be used as an antibody tool for researching PEDV virus infection and anti-infection mechanisms.

The invention discloses a method for purifying a toxoplasma gondii recombinant antigen, which comprises the following steps: selecting a full-length sequence of a toxoplasma gondii surface antigen SRS3, connecting the full-length sequence with a carrier through enzyme digestion to form plasmids, performing mass expression in escherichia coli, sequentially performing ultrasonication and centrifugation to collect cell supernatant, and performing deep filtration; the specific affinity chromatography column is coupled with an IgG subtype antibody obtained by hybridizing splenocytes and myeloma cells of a BALB / c mouse infected with toxoplasma gondii through a hybridoma technology. And through affinity chromatography purification, the toxoplasma gondii antigen protein with relatively high purity is finally obtained. According to the method, the toxoplasma gondii recombinant antigen can be stably and effectively purified from the cell supernatant. The method provided by the invention has the advantages of high extraction purity, simple process, capability of realizing one-step purification, stable antibody and the like, and has important significance for realizing purification of the toxoplasma gondii recombinant antigen.

The invention relates to a preparation method and application of hybridoma cells based on an electrofusion technology, the preparation method of the hybridoma cells based on the electrofusion technology comprises the following steps: taking splenocytes and myeloma cells of an antigen-immunized mouse for electrofusion, screening to obtain the hybridoma cells, and the antigen is serum amyloid protein A. The invention also relates to a preparation method of the hybridoma cells based on the electrofusion technology. The hybridoma cell capable of generating the serum amyloid protein A antibody is obtained in an electrofusion mode, the fusion efficiency of the hybridoma cell is improved, and the problem that the yield of the antibody is reduced due to the fact that the fusion efficiency of the hybridoma cell is low is solved.

The invention relates to a monoclonal antibody aiming at IGF2, a heavy chain variable region of the monoclonal antibody is shown as SEQ ID NO: 1, a light chain variable region of the monoclonal antibody is shown as SEQ ID NO: 2, and the invention also relates to application of the monoclonal antibody in detection of IGF2. According to the invention, antiserum aiming at IGF2 is obtained by immunizing a rat with an antigen, hybridoma cells are constructed by utilizing splenocytes of the rat, the monoclonal antibody with high affinity and specificity is successfully screened out, and the sequence of the antibody is obtained. The monoclonal antibody disclosed by the invention can be specifically combined with IGF2, is high in affinity and good in specificity, and can be used for detecting the IGF2 protein specifically expressed in adrenal tumor. Therefore, the compound can be used as an adrenal tumor diagnostic agent for early diagnosis of adrenal tumor.

The invention provides a monoclonal antibody capable of specifically recognizing a CV-A5 virus and having neutralizing activity and application of the monoclonal antibody. The monoclonal antibody is prepared from a CV-A5 virus antigen immunized mouse and splenocytes of the mouse through a cell fusion technology, and amino acid sequences of three CDR regions of a heavy chain variable region of the monoclonal antibody are sequentially shown as SEQ ID NO.1-3; the amino acid sequences of the three CDR regions of the light chain variable region are sequentially as shown in SEQ ID NO.4-6. The monoclonal antibody can be specifically combined with a CV-A5 virus, and is not combined with enteroviruses such as EV-A71, CV-A10, CV-A6, CV-A16 and the like. The monoclonal antibody targets conformational epitopes, has neutralizing activity, can specifically recognize CV-A5 viruses, is an ideal CV-A5 antigen detection antibody, and is beneficial to acceleration of the research and development process of hand-foot-mouth multivalent vaccines containing CV-A5 pathogens.

The present invention relates to a method for preparing a kelp extract exhibiting immune-enhancing activity and to the use thereof. The kelp extract according to the present invention has the effect of significantly increasing the expression of macrophage activity biomarkers COX-2 (Genome Name: PTGS2) and iNOS (Genome Name: NOS2), and immune-related cytokines TNF-α (Genome Name: TNF), IL-1β (Genome Name: IL1B), and IL-6 (Genome Name: IL6). In animal experiments, it increases natural killer (NK) cell activity, macrophage proliferation, macrophage phagocytosis, and splenocyte proliferation. Furthermore, compared to kelp extracts prepared by conventional methods, it exhibits significantly improved immune-enhancing efficacy and can be usefully utilized in pharmaceutical compositions for immune enhancement, health functional food compositions, etc.

The invention relates to a monoclonal antibody aiming at PGRMC1, a heavy chain variable region of the monoclonal antibody is as shown in SEQ ID NO: 1, a light chain variable region of the monoclonal antibody is as shown in SEQ ID NO: 2, and the invention also relates to application of the monoclonal antibody in detection of PGRMC1. According to the invention, antiserum aiming at PGRMC1 is obtained by immunizing a rat with an antigen, hybridoma cells are constructed by utilizing splenocytes of the rat, the monoclonal antibody with high affinity and specificity is successfully screened out, and the sequence of the antibody is obtained. The monoclonal antibody provided by the invention can be specifically combined with PGRMC1, has high affinity and good specificity, and can be used for detecting PGRMC1 protein specifically expressed in adrenocortical carcinoma. Therefore, the compound can be used as an adrenocortical cancer diagnostic agent for early diagnosis of adrenocortical cancer.

The invention relates to a Brucella BP26 protein, a preparation method thereof, and a preparation method and application of a monoclonal antibody of the Brucella BP26 protein. The amino acid sequence of the BP26 protein is shown as SEQ ID NO: 1; brucella BP26 protein is selected as an antigen, a homologous recombination technology is utilized to construct a BP26 prokaryotic expression vector, IPTG (isopropyl-beta-d-thiogalactoside) induced expression is carried out, and purification is carried out to obtain the recombinant BP26. An animal is immunized with the Brucella BP26 protein; fusing the tumor cells with splenocytes of an immunized animal, taking a cell supernatant culture solution to detect titer, and screening hybridoma cells; and collecting and purifying the monoclonal antibody secreted by the hybridoma cells. According to the invention, efficient soluble expression is realized by optimizing a BP26 gene sequence, and high-immunogenicity protein is obtained through purification; the protein is used for immunizing and screening a high-affinity monoclonal antibody. The BP26 protein and the monoclonal antibody can improve the Brucella detection sensitivity and specificity, are suitable for vaccine development, and have important clinical application value.

The invention provides a monoclonal antibody for sandwich detection of testosterone as well as a preparation method and application thereof, and the preparation method comprises the following steps: step S1, preparing testosterone-3-(O-carboxymethyl) oxime, and activating the testosterone-3-(O-carboxymethyl) oxime; taking a testosterone antibody P2G1 after activation, and dropwise adding the activated testosterone-3-(O-carboxymethyl) oxime according to the proportion for reaction to prepare an immunogen; s2, immunizing an animal based on the prepared immunogen; s3, fusing splenocytes and myeloma cells of the immune animals into hybridoma cells, and selecting real hybridoma cells; and S4, separating and purifying the antibody from the screened hybridoma cells. The invention provides a preparation method of a raw material and a reagent for testosterone sandwich method detection, solves the problems of insufficient sensitivity, narrow detection range and poor specificity of a micromolecule compound testosterone competition method reagent, and provides a new thought and method for rapid and accurate testosterone detection.

The invention provides a tortoise plastron peptide with immunomodulatory activity, and a preparation method, a product and application thereof, and relates to the field of biological medicines. According to the method, total protein in the tortoise plastron is extracted by means of decoction, concentration, extraction, filtration and the like, meanwhile, the content of fat-soluble impurities and mineral calcium in the extract is reduced, and then the total protein is subjected to enzymolysis to obtain polypeptide; the obtained polypeptide has good immunocompetence, and can effectively improve the multiplication capacity of splenocytes, promote macrophages to generate immune response and enhance the immune regulation function.

The invention discloses a monoclonal antibody, a hybridoma cell strain, a detection kit, a detection method and application, and relates to the technical field of animal epidemic disease prevention and control. According to the invention, a monoclonal antibody is obtained and is obtained by secreting a hybridoma cell strain DRV-4G3 with the preservation number of CGMCC (China General Microbiological Culture Collection Center) No.46567. The specific monoclonal antibody aiming at the duck reovirus muNS protein is prepared and obtained through an immune mouse and splenocyte fusion technology, the monoclonal antibody can recognize the muscovy duck reovirus and the novel reovirus at the same time, and the method for detecting the duck reovirus is established on the basis of the monoclonal antibody and can be used for large-scale serological detection. And reovirus vaccine immune antibody dynamic monitoring is achieved, and good development and practical application values are achieved.

The invention provides a universal diagnostic antibody for avian influenza virus as well as a preparation method and detection application thereof, and belongs to the technical field of biological detection. The invention provides a preparation method of an avian influenza virus universal diagnostic antibody. The preparation method comprises the following steps: intercepting an avian influenza virus M2 protein coding sequence, carrying out codon optimization, synthesizing by adopting an overlapping extension PCR (Polymerase Chain Reaction) technology, inserting pGEX-4T-1 plasmid, carrying out protein expression in E.coli Rosetta (DE3) competent cells, and purifying to obtain GST-M2 protein. GST-M2 protein is used for immunizing a mouse, after the titer of a serum antibody is detected, splenocytes are prepared and fused with SP2 / 0 cells, a strain of GST negative and GST-M2 positive hybridoma cells is screened out, and the mouse ascites monoclonal antibody is prepared after amplification culture. The M2 monoclonal antibody can specifically recognize M2 recombinant protein and can be used for detecting different avian influenza subtypes.

This invention discloses a method for screening TNFR1-targeting active ingredients in Tremella fuciformis fermentation broth, belonging to the field of active ingredient screening. The method includes: preparing and immobilizing a recombinant TNFR1 extracellular domain protein with a His tag; optimizing the conditions for preparing Tremella fuciformis fermentation broth and removing polysaccharides; subjecting the fermentation broth to affinity incubation with the immobilized protein to specifically capture the binding components; washing to remove impurities and then competitively eluting with imidazole phosphate buffer; finally, separating the TNFR1-targeting active ingredients by the methanol-chloroform method. This method utilizes the specific affinity of the TNFR1 target to efficiently and accurately enrich immunologically active ingredients from the complex Tremella fuciformis fermentation broth. Experiments have shown that the obtained components can significantly promote the proliferation of macrophages and splenocytes, providing a new source of raw materials and technical support for the development of novel immune-enhancing products.

The present invention relates to a composition for treating psoriasis diseases using nano-graphene oxide. It was confirmed that the nano-graphene oxide of the present invention regulates inflammatory immune cell subtypes and tissue-resident memory cells, which are psoriasis-pathogenic immune cells. In addition, in an acute psoriasis animal model, it was confirmed that skin thickness, scaling, and erythema were reduced, and the thickness of the epidermal layer was decreased. Furthermore, it was confirmed that immune cells are regulated by reducing psoriasis-pathogenic immune cells and increasing immunoregulatory cells in splenocytes. Moreover, it was confirmed that mitochondrial functions are regulated by increasing mitochondrial ROS and oxygen consumption rates in psoriasis-pathogenic immune cells.

The invention relates to the technical field of medicine in molecular biology and immunology, and discloses a porcine torovirus N protein monoclonal antibody as well as a preparation method and application thereof. A 6 * His tag is added into an optimized PToV N protein gene (the sequence is shown as SEQ ID NO.1), the gene is connected with a carrier to form a recombinant plasmid, the recombinant plasmid is converted into a competent cell to obtain a recombinant protein, the recombinant protein is purified and immunized with a mouse, splenocytes and myeloma cells SP2 / 0 of the mouse are collected for mixed culture, hybridoma cells capable of stably secreting a PToV-N protein monoclonal antibody are screened, and the PToV-N protein monoclonal antibody is obtained. And injecting the hybridoma cells into a mouse body to obtain the PToV-N protein monoclonal antibody. The screened monoclonal antibodies mAb 6D8 and mAb 7H3 are high in titer, high in sensitivity and high in specificity through WB and IFA detection, and can be used for preparing porcine torovirus detection kits, drugs and the like, and a research basis is provided for disease diagnosis, prevention and control of porcine torovirus.

Compositions for determining the efficacy and / or potency of a vaccine preparation are described herein. Splenocytes from immunized animals are isolated and can be frozen. Upon thawing such cells are activated by exposure to a series of dilutions of a vaccine preparation being tested and a series of dilutions of a reference vaccine with known characteristics. Cells secreting immunogen-specific antibody and cells secreting nonspecific antibody are enumerated, as is the amount of immunogen-specific and nonspecific antibody produced. Comparison between the results from the vaccine preparations provides a measure of relative vaccine efficacy and / or potency.

The invention relates to the technical field of protein peptide preparation, in particular to a protein peptide preparation for enhancing immunity and a preparation method thereof. The protein peptide preparation is prepared from the following components in parts by weight: 45 to 60 parts of bovine whey protein peptide, 8 to 15 parts of fistular onion seed protein peptide, 5 to 8 parts of 7-hydroxychromone derivative, 0.01 to 0.03 part of emulsifier and 0.02 to 0.05 part of stabilizer, the structural formula of the 7-hydroxychromone derivative is # imgabs0. Related biochemical index detection on a mouse immunocompromised model finds that through cooperative use of the fistular onion seed protein peptide, the 7-hydroxychromone derivative and the bovine whey protein peptide, the damage of immune organs of the immunocompromised mouse can be reduced; the division and proliferation of splenocytes are promoted to enhance the immune function of the body.

Described herein is an engineered viral subunit vaccine encoding ASFV capsid proteins (e.g., P72 and Penton) for use in pigs. The vaccine with either P72 or Penton tested in mice elicited significantly higher levels of antigen-specific IgM and IgG, increased frequency of antibody-secreting cells in the bone marrow, and higher quality germinal centers in the spleens of immunized mice. Enhanced T cell responses were also observed, with higher levels of IFN-γ and TNF-α in splenocytes. Immunogenicity assessments in pigs further supported these findings, with both MB-P72 and MB-PN180Q inducing robust B cell and T cell responses. These findings show that expression of capsid proteins P72 and Penton in membrane-bound forms via mRNA preserves their native multimeric structure and enhances their immunogenicity and provides a basis for an effective ASFV vaccine.

In some embodiments, the Disclosure provides a method for generating CAR T cells in situ. The Disclosure provides lipid nanoparticles that selectively target splenic cells, in particular lymphocytes, such as T cells. The lipid nanoparticles provided herein have an apparent pK of less than 6. a It contains a five-component composition that includes a permanent anionic lipid that imparts [a certain characteristic].