42 results about "Clostridium perfringens toxoid" patented technology

The invention discloses a monoclonal antibody blocking ELISA testing method of a clostridium perfringens alpha toxin, which is used for early testing of the alpha toxin and provides an effective testing tool for early diagnosis, spreading prevention and subsequent research of clostridium perfringens. The method comprises the following two steps: I, establishing the monoclonal antibody blocking ELISA testing method; and II, determining a result judgment criterion. The monoclonal antibody blocking ELISA testing method which has good specificity and stability and high sensitivity, is quick, simple and convenient, and tests the alpha toxin efficiently is established for the first time, and the method is simple to implement and high in testing speed and can achieve quantitative testing and massive testing. In a word, monoclonal antibody blocking ELISA is expected to play an important role in prevention and treatment of clostridium perfringens diseases.

The invention provides a bovine type-A clostridium perfringens subunit vaccine and a preparation method and an application thereof. The subunit vaccine comprises a bovine type-A clostridium perfringens alpha-toxin protein truncation body and a pharmacologically acceptable adjuvant. The bovine type-A clostridium perfringens alpha-toxin protein truncation body is an antigen determinant amino acid sequence from 255th to 372nd amino acids of alpha-toxin protein. The subunit vaccine has the following advantages that 1) safety problems due to uncompleted inactivation are not existed; 2) the qualityis controllable, and the difference between the batches is not existed; 3) the production equipment and space requirement is low, the expression level is high, and the cost is low; and 4) toxin dispersion risk is not existed, and the safety of operators can be guaranteed.

The invention discloses a clostridium perfringens alpha, beta 1, beta 2 and epsilon toxin coexpression vector; vectors are respectively pETDuet-1 and pRSFDuet-1 expressed in the same host bacteria, and the vectors are respectively connected with clostridium perfringens alpha and epsilon toxin encoding genes and clostridium perfringens beta 1 and beta 2 toxin encoding genes. On the basis, the invention also discloses a construction method and an expression method of the coexpression vector.

The invention discloses a humanized single-chain antibody 8B of clostridium perfringens alpha-toxin. The humanized single-chain antibody 8B is screened from a phage antibody library Source bioscience, is a full-humanized antibody of the clostridium perfringens alpha-toxin and can be used for achieving a toxin neutralization treating effect, overcoming various side effects of a heterologous antibody and eliminating complex steps and high cost of humanization modification on the heterologous antibody; and the single-chain antibody is small in molecular weight, strong in in-vivo penetrability and capable of rapidly reaching to damaged tissues and cells to exert an antitoxin effect, so that the double aims of economy and high efficiency are achieved. The alpha-toxin has a certain homology with other types of toxins of clostridium perfringens, so that the antibody drug possibly generates inhibiting and treating effects for other types of toxins and can also be used as a detecting reagent to detect the clostridium perfringens alpha-toxin.

The invention relates to a clostridium perfringen alpha toxin genetic engineering vaccine and application thereof. The clostridium perfringen alpha toxin genetic engineering vaccine is prepared by designing a specific primer, implementing polymerase chain reaction (PCR) amplification on A-type clostridium perfringen DNA to obtain an alpha toxin gene segment, cloning the gene segment onto a pMD18-T Vector, selecting a positive recon and linking the positive recon onto a PET-28 alpha (+) plasmid; introducing the recombinant plasmid to a receptor strain to successfully construct a recombinant strain BL21 (DE3)-alpha; emulsifying the prepared alpha toxin protein and an aluminum hydroxide adjuvant at a volume ratio of 9:1, and adding 0.01% thiomersal to obtain the clostridium perfringen alpha toxin genetic engineering vaccine. The clostridium perfringen alpha toxin genetic engineering vaccine is easy for realization of industrial production, simple to operate and good in safety, and is capable of effectively preventing such diseases as animal necrotic enteritis, enterotoxemia, and human and animal traumatic gas gangrene caused by the A-type clostridium perfringens.

The invention provides a clostridium perfringen alpha toxin genetic engineering vaccine and a preparation method thereof. Compared with wild type alpha toxins, clostridium perfringen alpha toxin recombination protein is characterized in that the 176th site histidine of an amino acid sequence is mutated to obtain asparagine. The clostridium perfringen alpha toxin genetic engineering vaccine is prepared from detoxifcation clostridium perfringen alpha toxin recombination protein of self-induction secretory expression. The vaccine has the advantages of being safer, better in immune efficacy, simpler in technology, lower in cost and the like, and can effectively solve the problems that in the prior art, the production technology of the vaccine is complexer, and the cost is higher.

The invention discloses a humanized single-chain antibody 7D of clostridium perfringens alpha-toxin. The humanized single-chain antibody 7D is screened from a phage antibody library Source bioscience, is a full-humanized antibody of the clostridium perfringens alpha-toxin and can be used for achieving a toxin neutralization treating effect, overcoming various side effects of a heterologous antibody and eliminating complex steps and high cost of humanization modification on the heterologous antibody; and the single-chain antibody is small in molecular weight, strong in in-vivo penetrability and capable of rapidly reaching to damaged tissues and cells to exert an antitoxin effect, so that the double aims of economy and high efficiency are achieved. The alpha-toxin has a certain homology with other types of toxins of clostridium perfringens, so that the antibody drug possibly generates inhibiting and treating effects for other types of toxins and can also be used as a detecting reagent to detect the clostridium perfringens alpha-toxin.

The invention relates to surface display C type clostridium perfringens alpha and beta toxin protein recombinant lactobacillus plantarum. The C type clostridium perfringens alpha toxin gene sequence,beta1 toxin gene sequence and beta2 toxin gene sequence are inserted in a genome of lactobacillus plantarum. The invention further relates to an establishment method and application of the recombinantlactobacillus plantarum. Through a PCR method, alpha, beta1 and beta2 toxin genes are successfully cloned on the whole genome of C type clostridium perfringens (CVCC 1158), a corresponding recombinant plasmid is established and electrically transferred into lactobacillus plantarum NC8, three strains of recombinant lactobacillus plantarum are obtained after induction expression, the oral mode is selected for immunizing a rat, the immune effect of recombinant bacteria is evaluated, the result shows that when the rat is immunized by orally taking the recombinant lactobacillus plantarum, the ratbody can be stimulated to generate humoral immunity and can also be stimulated to generate cellular immunity, and a good protection effect on an animal body is realized.

The invention relates to the field of animal bacteriology and provides a capture ELISA (enzyme-linked immunosorbent assay) detection method for a Clostridium perfringens Alpha-toxin antibody; in the method, prepared type-A Clostridium perfringens exotoxin is used as an antigen that is used for immunizing a rabbit to obtain rabbit serum and for acting as a binding antigen for capture ELISA after being concentrated and purified; the detection method comprises the steps of (1) enveloping; (2) sealing; (3) adding an antigen; (4) adding a sample to be tested; (5) adding a secondary antibody; (6) developing color; (7) ending; (8) judging results. The detection method has the advantages of high specificity, high sensitivity, good repeatability, low cost and the like; in addition, the detection method is suitable for detecting samples in large batch, and an effective and simple serological diagnostic method is provided for the detection and research on such toxin.

The present disclosure relates to nanoparticle compositions for use as vaccines against Clostridium perfringens in poultry which causes necrotic enteritis in poultry. Such compositions include one or more Clostridium perfringens extracellular proteins entrapped in a polyanhydride or chitosan nanoparticle. The one or more Clostridium perfringens extracellular proteins may include one or more Clostridium perfringens toxins, such as, for example, alpha toxin (CPA), beta toxin (CPB), epsilon toxin (ETX), iota toxin (ITX), perfringolysin O (PFO), enterotoxin (CPE), beta2 toxin (CPB2), or NetB toxin. In some aspects, the composition further includes a Salmonella enteritidis flagellar protein. The present invention also includes methods for the oral delivery of one or more Clostridium perfringens extracellular proteins to the mucosal membrane of the intestinal tract of a bird of the order Galliformes.

The invention provides clostridium perfringens beta toxin mutant protein as well as a preparation method, application and a vaccine thereof, and relates to the field of biological products. The clostridium perfringens beta toxin mutant protein is prepared by mutating 212th arginine and 266th tyrosine of wild clostridium perfringens beta toxin with an amino acid sequence as shown in SEQ ID NO.3 into alanine, and the toxicity of the clostridium perfringens beta toxin mutant protein is remarkably reduced compared with that of the wild clostridium perfringens beta toxin. Meanwhile, good immunogenicity is also reserved.

The invention provides a clostridium perfringens epsilon toxin mutant protein as well as a preparation method, application and a vaccine thereof, and relates to the technical field of biology. The clostridium perfringens epsilon toxin mutant protein includes alanine mutated by 151-position amino acid of clostridium perfringens epsilon toxin shown in an amino acid sequence SEQ ID NO.3. Compared with a wild type, the clostridium perfringens epsilon toxin protein has significantly reduced toxicity and good immunogenicity. The vaccine can alleviate technical problems such as complex production process, high cost, complex antigen, inter-batch difference and large side effects of inactivated vaccine.

The invention discloses a Clostridium perfringens alpha toxin visual detection method. A smartphone image processing technology and silicon dioxide microsphere antibody coupling are combined to detectfood-borne clostridium perfringens alpha toxin (CPA). According to a double-antibody sandwich principle, the prepared CPA polyclonal antibody is coupled with silicon dioxide microspheres, CPA fluorescence detection is performed through antibody-labeled FITC fluorescence, and finally, the CPA content is detected in real time in combination with an image processing technology of a smartphone APP. Compared with a traditional clostridial toxin detection method, the nano-microsphere combined intelligent mobile phone CPA detection system is a more sensitive, stable and convenient detection method,and a new method is provided for detection of natural CPA in food.

The invention discloses a fusion protein vaccine for chicken necrotic enteritis and a preparation method thereof. The fusion protein vaccine is prepared by connecting Clostridium perfringens NetB toxin, Tpel toxin with the C tail end and the C end cut off of alpha toxin in series through a joint segment A (EAAAK) 4A and expressing and purifying the toxin in escherichia coli BL21(DE3). The fusion protein and a propolis adjuvant are mixed in an equal volume mode to immunize broiler chickens, and the result shows that the body can be induced to generate a high antibody level, and infection of G-type Clostridium perfringens can be remarkably resisted. The fusion protein vaccine can keep the space structure and immunogenicity of three toxin proteins, can avoid the complex process and biosafety problems of a traditional vaccine, is easy to produce industrially, has the advantages of being low in preparation cost, high in safety and excellent in efficacy, and can well prevent chicken necrotic enteritis clinically.