52 results about "Complete adjuvant" patented technology

The invention provides a method for preparing triple vaccine for preventing porcine transmissible gastroenteritis, porcine epidemic diarrhea and porcine rotavirus. The method comprises the following steps: inoculating a host-cell line with a 90 percent grown monostratum against a porcine transmissible gastroenteritis virus, a porcine epidemic diarrhea virus and a porcine rotavirus respectively, and adding a cell maintenance media into the host-cell lines respectively to be cultured at 37 DEG C; after cytopathic effect reaches over 75 percent, collecting viruses to be stored at 20 DEG C below zero for standby; mixing the viruses according to 10 TCID50 in 1:1:1, and simultaneously adding Freund's complete adjuvant and immunopotentiator into the mixture to inactivate the mixture by formaldehyde at 37 DEG C for 24 hours; and adding an oil adjuvant into the mixture to prepare a vaccine of water-oil-water preparation. The method can be used for preparing the triple vaccine for preventing the porcine transmissible gastroenteritis, the porcine epidemic diarrhea and the porcine rotavirus so as to solve the problem that the diseases do not have an effective medicine to treat currently.

The invention discloses a preparation method of a diethylstilbestrol antibody. The preparation method of the diethylstilbestrol antibody comprises the following steps of: (1) performing a reaction of diethylstilbestrol and 4-bromine butyric acid hexylester, exposing a carboxyl group by saponifying and hydrolyzing, and synthesizing diethylstilbestrol artificial haptene; (2) performing a reaction of the diethylstilbestrol artificial haptene and bovine serum albumin by a mixed anhydride method to obtain diethylstilbestrol artificial immunogen; and (3) emulsifying the diethylstilbestrol artificial immunogen and Freund's complete adjuvant (initial immunization) or Freund's incomplete adjuvant, subcutaneously injecting a rabbit, and separating blood serum when the antibody titer is stable to obtain the diethylstilbestrol antibody. The preparation method of the diethylstilbestrol antibody has the advantages of simpleness, convenience and low cost; and the prepared diethylstilbestrol antibody has high specificity, high antibody titer and indirect ELISA experimental antibody titer of over 1 million, can reduce detection cost when used for immunologically detecting diethylstilbestrol, and is easy to promote and use.

The invention provides preparation methods for duck hepatitis virus immunogen and hyperimmune serum and application of the duck hepatitis virus hyperimmune serum. According to the preparation methods and the application of the duck hepatitis virus hyperimmune serum, the duck hepatitis virus immunogen is obtained through inoculating a serum 1 type duck hepatitis virus CH60 strain DHAV-1 (Duck Hepatitis A Virus type 1) or a serum 3 type duck hepatitis virus CH1 strain DHAV-3 (Duck Hepatitis A Virus type 3) to an allantoic cavity of a chick embryo of 9-10 days old or a duck embryo of 10-12 days old and carrying out proliferation and treatment, and the hyperimmune serum is obtained through mixing the duck hepatitis virus immunogen with a Freund's complete adjuvant or Freund's incomplete adjuvant to prepare solutions of different concentrations, carrying out repeated immunization on immune animals and then sampling and collecting blood and can be applied to the diagnosis and detection on a duck hepatitis virus. The preparation methods provided by the invention have the advantages that the conditional requirements are low, the operation is simple, and the obtained immunogen can meet the requirements on the preparation of specific antiserum.

The invention discloses a soft-shelled turtle systemic septicemia spherical virus inactivated vaccine and its preparation method, and is characterized in that the vaccine contains the inactivated soft-shelled turtle systemic septicemia spherical virus with the collection number being CGMCCNo.5378. The preparation method comprises the following steps of: cutting internal organ of sick soft-shelledturtles into pieces, adding into a TNE buffer, centrifuging, taking a supernatant, centrifuging, taking a white precipitate, suspending in the TNE buffer, centrifuging the precipitate, taking the supernatant, and centrifuging to obtain a white precipitate, namely the purified virus; diluting the purified virus by the use of aseptic normal saline until the final concentration of viral protein reaches 0.5-1mg / ml, and adding 0.5% (final volume) of formalin for inactivation in a thermostat water bath cauldron; and completely emulsifying inactivated virus and a complete Freund's adjuvant accordingto the volume ratio of 1:1 to obtain the inactivated vaccine. The soft-shelled turtle systemic septicemia spherical virus inactivated vaccine proposed for the first time is safe and effective and hasstrong immunity, and the preparation method is simple and easy to operate.

The invention provides an immunogen for obtaining an Nrf1D protein antibody, the Nrf1D protein antibody and an Elisa detection kit. The immunogen is obtained as follows: polypeptide with the amino acid sequence shown in SEQ ID NO.1 is synthesized firstly, Sulfo-SMCC is used as a coupling agent, the polypeptide and KLH (keyhole limpet hemocyanin) are coupled; the antibody is obtained as follows: immunogen is taken as an antigen, the antigen is emulsified with FCA (freund's complete adjuvant) to immunize a New Zealand white rabbit, blood of the immunized New Zealand white rabbit is centrifuged, and a supernatant after centrifugation is collected; the Elisa detection kit comprises components including an ELISA plate, an enveloping solution, a blocking solution, an ELISA secondary antibody, a TMB color developing solution, a stop solution, a detection antigen and the Nrf1D protein antibody, the detection antigen is the polypeptide with the amino acid sequence shown in SEQ ID NO.1, glutaradehyde is used as a coupling agent, and the polypeptide and BSA bovine serum albumin are coupled.

The invention discloses a preparation method of a monoclonal antibody to chloramphenicol (CAP). The method comprises the following steps of: (1) preparing an antigen of CAP-BSA by a diazotization method; (2) conducting lymph immunization to mice with the CAP-BSA antigen, leaving the antigen and a Freund's complete adjuvant to complete emulsification during the first time immunization, and during the second time and the third time immunization, leaving the antigen and a Freund's incomplete adjuvant to complete emulsification, and keeping an interval of 7 days between each time, and 3 days before fusion, carrying out direct antigen injection to the abdominal cavity to booster immunization; (3) measuring serum titers of the mice by an enzyme-linked immunosorbent assay (ELISA) method; (4) selecting a mouse with the highest serum titer, and taking a spleen cell and a myeloma cell for in vitro fusion; (5) culturing and screening a fusion cell in a selective medium, conducting positive cell detection and screening for cloning culture, further performing positive cell cloning and screening for positive cloning, expanding culture and cryopreservation; (6) injecting a cell strain undergoing expanded culture into the abdominal cavities of the mouse so as to produce a lot of ascites; (7) using a chromatographic column to purify the monoclonal antibody against chloramphenicol in the ascites. The prepared monoclonal antibody against CAP-BSA has the advantages of high sensitivity, strong specificity and good practicality.

The invention discloses a capparis acutifolia sweet exact and a preparation method and an application thereof. The capparis acutifolia sweet extract is obtained by taking roots and stems of plant-capparis acutifolia sweet, smashing, then carrying out heating reflux and cold soaking or leakage for extraction, carrying out vacuum concentration for obtaining a crude extract and using chloroform for extracting the crude extract, and can be applied in the preparation of drugs for treating neuropathic pain, treating inflammatory pain, treating cancer pain, treating diabetic pain, treating rheumaticpain, treating traumatic injury pain, treating sore throat, treating toothache or treating abdominal pain, and the capparis acutifolia sweet extract has great analgesic and anti-inflammation activityand obvious effect for Freund's complete adjuvant arthritis. The preparation process is characterized by low cost and easy realization of industrial production.

The invention relates to a recombinant Wzt protein rabbit serum polyclonal antibody and a preparation method thereof, which relate to the field of antibody preparation, and fill the gap of applying recombinant Wzt protein for preparing the specific polyclonal antibody. The preparation method of the polyclonal antibody comprises the steps of after mixing and emulsifying the recombinant Wzt protein and an equivoluminal freund's complete adjuvant, carrying out multiple sites subcutaneous injection on the back parts of female rabbits; carrying out secondary immunizing: after one week, after mixing and emulsifying the recombinant Wzt protein and the equivoluminal freund's incomplete adjuvant, carrying out multiple sites subcutaneous injection on the back parts of the female rabbits; carrying out thrice immunizing and quartic immunizing every other week according to a secondary immunizing process; collecting venous blood of female domestic rabbit ears after immunizing each time, and detecting a rabbit serum antibody titer; at one week after quartic immunizing, collecting rabbit whole blood, centrifugally collecting a supernatant to obtain a rabbit serum, and obtaining the recombinant Wzt protein rabbit serum polyclonal antibody after purifying. The recombinant Wzt protein rabbit serum polyclonal antibody has favorable specificity, and can be used for detecting Brucella Wzt protein.

The invention relates to a method for preparing antibody used to check lean meat, wherein it comprises: collecting the animal blood and separating serum to prepare full-serum protein solution; the mixing the solution into carbonate baffle liquid; then adding azo liquid, to prepare carrier couple material; adding the carrier couple material and Freund's complete adjuvant into penicillium notatum bottle, vibrating it on mixer to be white color, to form CL Freund's complete adjuvant emulsion liquid; using emulsifying another carrier couple material and Freund's incomplete adjuvant; and planting the prepared CL Freund's complete adjuvant emulsion liquid on animal to be used in immunization; then separating serum to obtain antibody. The invention uses itself protein, especially the full-serum as the immunization carrier of CL, to make the immunity animal generate special antibody whose ELISA effect can reach more than 40thousand; and the immunity gold testing method built from said antibody can quickly test CL of urine sample, while its sensitivity is 40ng / ml.

A preparation method of a grass carp ATG12 polyclonal antibody comprises the steps: firstly, taking grass carp liver RNA as a template, carrying out reverse transcription to synthesize cDNA, then taking the cDNA as a template, amplifying by PCR with primers and recovering a target fragment, and then constructing an ATG12 / pGEX-4T-1 prokaryotic expression vector; transferring the ATG12 / pGEX-4T-1 prokaryotic expression vector into competent cells of escherichia coli DH5[alpha], treating, then carrying out ultrasonic crushing and centrifugation to obtain a supernatant and a precipitate, and then purifying the precipitate to obtain purified ATG12 fusion protein; and finally, fully mixing and emulsifying the purified ATG12 fusion protein as an antigen with a Freund's complete adjuvant with the same volume with the ratio of 1:1, immunizing pure New Zealand white rabbits, extracting heart blood, and separating serum, to obtain the ATG12 polyclonal antibody. The antibody has the advantages of high specificity, low cost and short period, and lays the foundation for obtaining the commercially available ATG12 antibody.

The invention discloses a recombined subunit vaccine of haemaphysalis concinna and a preparation method thereof. The recombined subunit vaccine is formed by mixing antigenic gene recombined protein of the haemaphysalis concinna and Freund's complete adjuvant (FCA), wherein the content of the antigenic gene recombined protein is 50 mg / ml, that is 50 mL rHc-23 and 950 ml Freund's complete adjuvant are mixed to prepare the recombined subunit vaccine of the haemaphysalis concinna; the amino acid sequence of the antigenic gene recombined protein is shown in the table SEQID NO:1; and the nucleotide sequence of the antigenic gene is shown in the table SEQID NO:2. Through screening and cloning, prokaryotic expression and separation and purification of protective antigen gene of the haemaphysalis concinna and the application effect test of the recombined subunit vaccine, the method shows that an expression product of recombined vector bacteria can be identified by rabbit anti-haemaphysalis concinna positive serum. In an animal immunity test, after rHc-23-FCA is subjected to three immune rabbits, the blood saturation rates of haemaphysalis larva, haemaphysalis middle and haemaphysalis imago are 40.3 percent, 45.6 percent and 41.3 percent respectively; and compared with the blood saturation rates of the haemaphysalis larva, haemaphysalis middle and haemaphysalis imago in a contrast set: 90.1 percent, 94.3 percent and 97.7 percent, the differences are remarkable. The method promotes the development of anti-haemaphysalis immunity, haemaphysalis control and haemaphysalis disease spread work.

The invention discloses an egg yolk antibody and a preparation method and device thereof. A test object is used for performing polypide reinvigoration and proliferation on tender eimeriida, giant eimeriida and poisoned eimeriida, after the insects are mixed in equal proportions, antigen is prepared by ultrasonic disruption, the obtained antigen solution is mixed with an equal volume of freund's complete adjuvant and freund's incomplete adjuvant separately, laying hens are immunized three times, egg yolk antibodies in collected egg yolk after three times of immunizationare extracted, purified and identified, and the antibody titer is evaluated; the research on different yolk powder preparation processes is carried out, the effect of lyophilized powder preparation is significantly higher than that of powder spraying technology, the matchingrelationship between the yolk and the dilution, the lyophilization temperature curve and other parameter conditions in the process are prepared, the cost is relatively low, the preparation efficiency is high, antibody loss rate is only 10%, high antibody titer, antibody titer after high-temperature powder spraying shows a high downward trend, and the method is obviously superior to the high-temperature powder spraying technology for preparing yolk powder.

The invention discloses a method for preparing serum for resisting Muscovy duck parvovirus, which comprises the following steps of: preparing the Muscovy duck parvovirus; preparing a primary immune Muscovy duck parvovirus antigen; preparing a secondary immune Muscovy duck parvovirus antigen; preparing a Freund's incomplete adjuvant vaccine for the Muscovy duck parvovirus from the secondary immuneMuscovy duck parvovirus antigen; preparing a Freund's complete adjuvant vaccine for the Muscovy duck parvovirus from the primary immune Muscovy duck parvovirus antigen; and immunizing the Muscovy duck by using the prepared antigens and vaccines to obtain the serum for resisting the Muscovy duck parvovirus. The serum prepared by the method has high titer, and has obvious effect of preventing and controlling the Muscovy duck parvovirus; the prevention rate of the serum is 96.6 percent; and the cure rate of the serum is about 70 percent.

The invention relates to the technical field of biological protein molecules, in particular to a kenaf mitochondrial protein COX3 antigen polypeptide and a method for preparing a polyclonal antibody and an application. An amino acid sequence of the antigen polypeptide is shown in SEQ ID NO: 2. The method includes coupling the antigen polypeptide with carrier protein KLH to obtain polypeptide-KLH coupling composite protein; mixing with a Freund's complete adjuvant with equal volume, emulsifying, and immunizing animals; collecting animal blood serum, collecting animal blood by carotid artery bloodletting when the titer reaches more than 1: 20000, and preparing antibody serum; and purifying the antibody serum by affinity chromatography to obtain the polyclonal antibody of the kenaf mitochondrial protein COX3. By synthesizing the kenaf COX3 polyclonal antibody, the expression difference of the kenaf COX3 protein in different cytoplasm of kenaf is detected, so as to determine the importantinfluence of COX3 on the development of kenaf anthers.

The invention provides a preparation method of micro-encapsulation coated yolk source cholecystokinin (CCK) antibodies. The preparation method comprises the following steps: preparing an antigens complex through adding freund's complete adjuvant in the mixture formed after a sheep derived CCK and BSA (Bovine Serum Albumin) are crosslinked to immunize laying hens, collecting eggs and obtaining the yolk derived CCK antibodies from the eggs, taking sodium alginate aqueous solution to perform the homogeneous emulsification with yolk derived CCK antibody liquid, dripping the emulsified solution into encystation solution consisting of chitosan and CaCl2 to be coated. The coated yolk derived cholecystokinin antibodies protect the effective actives of the yolk derived cholecystokinin antibodies, and have enteric solubility. According to the preparation method, a micro-encapsulation technology is adopted, and chitosan and sodium alginate are used as the coating materials which are natural polysaccharide, are low in cost, are taken orally and have no any toxic and side effect.

The invention discloses a rabbit alliin antibody acquired through the immune response of the alliin antigen in the technical field of biological preparations. The rabbit alliin antibody contains the alliin antigen and a Freund's complete adjuvant, the alliin antigen and the Freund's complete adjuvant with equal amount are mixed to obtain an adjuvant antigen, the alliin antigen is injected into thebody of a rabbit, and the alliin antibody is obtained after the immune response. According to the alliin antigen and the rabbit alliin antibody, the immunogenicity of alliin is solved, the antibody is acquired through the immunization of the rabbit, and the alliin antibody can be applied to research of the concentration detection, in-vivo distribution and pharmacological action mechanisms of alliin and has important application values in the foundation and clinical researches of alliin.

The invention discloses a rabbit monoclonal antibody preparation method. The method comprises the following steps of 1, preparing immune serum; 2, affine purifying an antibody; 3, obtaining a mass spectrum; 4, obtaining an antibody variable region gene spectrum; 5, obtaining the monoclonal antibody, wherein, in step 1, heteromorphic rabbits with bas mutations and wild parents b9 are selected, subcutaneous inoculation of freund's complete adjuvant on the rabbits, immunization is performed every three days, immunization is performed three times, injection is performed one time two weeks after immunization, one month after immunization, NG-XMTTM protein is subjected to intravenous injection at the tails of the rabbits, a sterile small test tube is used for collecting immunized rabbit blood, after marking, the rabbits are delivered to a preparation room, unimmunized rat serum is adopted as control, ELISA is adopted for determining antibody valence, the ELISA is used for determining the OD490 light absorption value quantification and qualitatively judging whether or not the immune serum is positive, and the positive immune serum is selected; the method is more efficient and rapider andhas high affinity.

The invention provides a preparation method for a monoclonal antibody of riemerella anatipestifer. The method comprises the following specific steps: (1) expanding the culture of the riemerella anatipestifer, inactivating formaldehyde, and then preparing suspension by using PBS to complete antigen preparation; (2) mixing the newly prepared antigen with a freund's complete adjuvant uniformly and stirring for 4 hours until a mixture is emulsified, then performing subcutaneous injection on mice, mixing the newly prepared antigen with the freund's incomplete adjuvant uniformly and stirring for 4 huntil a mixture is emulsified after two weeks of immunization, performing subcutaneous injection on the mice again to obtain immunized mice after one week of immunization, and detecting antibody titer is 1 to 6400; (3) performing cell fusion on the spleen cells of the immunized mice and SP2 / 0 cells; 4) performing positive screening and cloning on fusion cells in step (3). According to the method,the antibody titer is increased by improving the immune effect, so that the immunization times are reduced, thereby achieving the effects of saving time and reducing materials.

The invention discloses a preparation method of a composite IgY antibody against common cold virus, streptococcus pneumonia and staphylococcus. The preparation method comprises the following steps: mixing the common cold virus, streptococcus pneumonia and staphylococcus as antigens to obtain Freund's complete adjuvant immunogen and Freund's incomplete adjuvant immunogen; performing subcutaneous and intramuscular immunization of a laying hen; extracting the IgY antibody from the egg yolk of the immunized hen by a water dilution method; and purifying the IgY antibody by a two-step ammonium sulfate salt fractionation method to obtain a composite IgY antibody against the common cold virus, streptococcus pneumonia and staphylococcus. A product prepared from the composite IgY antibody with specific resistance against the common cold virus, streptococcus pneumonia and staphylococcus can be used for controlling the upper respiratory infection inflammation such as common cold, rhinitis, sinusitis, tonsillitis, sphagitis and bronchitis caused by the common cold virus, streptococcus pneumonia and staphylococcus and the viruses and bacteria with the same antigens.

The invention discloses a kanamycin antibody and a preparation process thereof. The kanamycin antibody includes, by weight, 58-80 parts of kanamycin sulfate, 5-25 parts of bovine serum albumin, 3-5 parts of egg albumin, 3-5 parts of Freund's complete adjuvant, 3-5 parts of Freund's incomplete adjuvant, 3-5 parts of carbodiimide, 1-2 parts of glutaraldehyde, 1-3 parts of goat-anti-mouse enzyme labeled secondary antibody, 2-3 parts of disodium hydrogen phosphate, 8-10 parts of sodium acetate, 15-20 parts of sodium dodecyl sulfate, 10-15 parts of Coomassie brilliant blue R-250, 8-12 parts of urea peroxide, 6-11 parts of ethanol, 5-7 parts of formaldehyde, 1-3 parts of a phosphate buffer solution, 1-3 parts of a washing solution PBST, and 1-3 parts of a sealing solution 5% pig serum. With the kanamycin sulfate and the bovine serum albumin as main raw materials, the antibody which can be competitively bond to and can be coated by the enzyme labeled antigen is prepared in a conjugation manner. The whole reaction process is simple, high-effective and economical and is excellent in availability.

The invention relates to cloning expression and polyclonal antibody preparation of black-headed gull IFN alpha protein and belongs to the technical field of bioengineering. The invention provides cloning expression and polyclonal antibody preparation of the black-headed gull IFN alpha protein. Part of fragments of the black-headed gull alpha interferon gene are cloned, a prokaryotic expression recombinant vector is constructed, recombinant protein is obtained to prepare a polyclonal antibody, a theoretical basis is provided for later researches on the antiviral activity of the polyclonal antibody and the antiviral mechanism of the black-headed gull alpha interferon gene, a new path is developed for prevention and control of epidemic diseases of black-headed gulls, and veterinary public health is deeply affected. When rabbits are immunized, a Freund's complete adjuvant and an incomplete adjuvant are selectively applied to be matched with protein injection, and the polyclonal antibody can be efficiently and rapidly prepared. The agar diffusion test proves that the recombinant protein of partial segment of the black-headed gull IFN alpha gene has favorable immunogenicity, and the titer of the antibody can reach 1: 8. The method can be well used for subsequent researches on the antiviral mechanism of the black-headed gull IFN alpha protein.

The invention discloses a preparation method of a mice antiserum resisting a toad skin transgelin-2 recombinant protein and belongs to the technical field of biological medicine. The preparation method is characterized by comprising the following steps of 1, preparing a toad skin transgelin-2 recombinant protein and a Freund's complete adjuvant into an agent A, and preparing the toad skin transgelin-2 recombinant protein and a Freund's incomplete adjuvant into an agent B, 2, carrying out mice hypodermic multipoint injection of the agent A, carrying out normal feeding for 12-16 days, carrying out hypodermic injection of the agent B, and then carrying out injection of the agent B every 12-16 days, wherein the agent B injection process is totally carried out 3-5 times, and 3, collecting heart blood of the treated mice and carrying out centrifugation to obtain a supernatant which is the mice antiserum. The preparation method can be used for handling and forecasting specific events of transgelin-2 antitumor research and has effects of prompting and marking transgelin-2. The mice antiserum can be obtained simply, has a low cost and has obvious effects.

The invention provides a preparation method and application of a candidate outer membrane protein of Aeromonas hydrophila vaccine, and designs outer membrane protein gene cloning primers; through prokaryotic recombination, using in vitro PCR amplification technology, amplifies target gene fragments, and constructs pET32a prokaryotic Express the recombinant plasmid of the vector, transform the recombinant plasmid to BL21 competent, and obtain a high-expression strain; induce the high expression of OMP P5 protein by IPTG inducer and purify the expressed protein by Ni-NTA resin column; the purified outer membrane protein and Freund's Insulin The complete adjuvant is completely emulsified at a volume ratio of 1:1, and the obtained sample is used as the vaccine protein of the Aeromonas hydrophila vaccine. In the present invention, the Aeromonas hydrophila subunit vaccine (outer membrane protein) can be used to obtain an immune protection rate of more than 90%. Compared with traditional vaccines, the subunit vaccine has low toxic and side effects, strong immunogenicity and long duration, High antibody titer and other advantages.

The invention discloses a preparation method and application of an anti-herpes simplex virus complex IgY antibody, and belongs to the technical field of herpes simplex virus resisting. The preparationmethod includes the following steps that step 1, an immunogen is prepared, specifically, a herpes simplex virus type 1 and a herpes simplex virus type 2 are taken and mixed and then the mixture is mixed with a Freund's complete adjuvant and a Freund's incomplete adjuvant separately and stirred and emulsified to prepare a water-in-oil Freund's complete adjuvant and Freund's incomplete adjuvant herpes simplex virus immunogen; and step 2, immunity is performed, specifically, the Freund's complete adjuvant immunogen is injected into laying hens. Application to ocular conjunctiva, a nasal cavity and an oral cavity is achieved, no limitations and side effects of systemic medication exist, the local application taking effect is quick, the dosage is low, the curative effect is good, the normal micro-ecological environment on the surface of the conjunctiva, the nasal cavity and the oral cavity is not absorbed and damaged, no toxic and side effects exist, and diseases of herpes labialis, dermatitis herpetiformis and herpes zoster caused by the herpes simplex virus type 1 and the herpes simplex virus type 2 and viruses with a common antigen with the herpes simplex virus type 1 and the herpessimplex virus type 2 are prevented and treated.

The invention discloses a doxycycline broad-spectrum monoclonal antibody and a preparation technology. The doxycycline broad-spectrum monoclonal antibody is prepared from the following raw materials in parts by weight: 40-52 parts of doxycycline, 4-20 parts of tetracycline, 2-6 parts of minocyclin, 2-4 parts of metacycline, 3-5 parts of bovine serum albumin, 5-8 parts of a Freund's complete adjuvant, 1-2 parts of Freund's incomplete adjuvant, 2-3 parts of HRP-Goat anti Mouse IgG, 3-6 parts of ammonium sulphate, 9-10 parts of disodium hydrogen phosphate, 16-20 parts of potassium dihydrogen phosphate, 12-16 parts of citric acid, 8-12 parts of ammonium dihydrogen phosphate, 9-15 parts of anhydrous ethanol, 1-3 parts of isobutyl chlorocarbonate, 2-4 parts of a coating buffered solution, 5-7 parts of confining liquid, 3-4 parts of a washing buffered solution, 6-10 parts of substrates and 20-25 parts of cell culture fluid, wherein the doxycycline and 4-Aminophenylacetic acid are used as main raw materials, and a diazo method is used for preparing the doxycycline broad-spectrum monoclonal antibody having antisera specificity. The doxycycline broad-spectrum monoclonal antibody has excellent biological properties, and is good in antisera titer and rejection capability.

The invention provides an immunogen for obtaining an Nrf1D protein antibody, the Nrf1D protein antibody and an Elisa detection kit. The immunogen is obtained as follows: polypeptide with the amino acid sequence shown in SEQ ID NO.1 is synthesized firstly, Sulfo-SMCC is used as a coupling agent, the polypeptide and KLH (keyhole limpet hemocyanin) are coupled; the antibody is obtained as follows: immunogen is taken as an antigen, the antigen is emulsified with FCA (freund's complete adjuvant) to immunize a New Zealand white rabbit, blood of the immunized New Zealand white rabbit is centrifuged, and a supernatant after centrifugation is collected; the Elisa detection kit comprises components including an ELISA plate, an enveloping solution, a blocking solution, an ELISA secondary antibody, a TMB color developing solution, a stop solution, a detection antigen and the Nrf1D protein antibody, the detection antigen is the polypeptide with the amino acid sequence shown in SEQ ID NO.1, glutaradehyde is used as a coupling agent, and the polypeptide and BSA bovine serum albumin are coupled.

The invention discloses a preparation method of a composite IgY antigen against propionibacterium acnes and staphylococcus and an application of the composite IgY antigen, and belongs to the technicalfield of resistance against propionibacterium acnes and staphylococcus. The preparation method comprises the following steps: step 1, preparation of immunogens: mixing propionibacterium acnes and fragments thereof with staphylococcus aureus, staphylococcus albus and fragments thereof, and then mixing with a Freund's complete adjuvant and a Freund's incomplete adjuvant respectively, conducting emulsifying by stirring to obtain the water-in-oil Freund's complete adjuvant and Freund's incomplete adjuvant immunogens against propionibacterium acnes and staphylococcus; step 2, immunization: injecting a Freund's complete adjuvant immunogen to laying hens. The specific composite IgY antigen against propionibacterium acnes and staphylococcus prepared by the invention is applied locally on face skin and whole body skin, and has advantages of rapid action, less dosage, good curative effect and no damage on normal micro-ecological environments of mucosa and skin without limitations and side effects of systemic administration.