+

WO1990007525A1 - Cytotoxine de la keratoconjonctivite et ses procedes d'utilisation et de production - Google Patents

Cytotoxine de la keratoconjonctivite et ses procedes d'utilisation et de production Download PDF

Info

Publication number
WO1990007525A1
WO1990007525A1 PCT/US1990/000106 US9000106W WO9007525A1 WO 1990007525 A1 WO1990007525 A1 WO 1990007525A1 US 9000106 W US9000106 W US 9000106W WO 9007525 A1 WO9007525 A1 WO 9007525A1
Authority
WO
WIPO (PCT)
Prior art keywords
cytotoxin
bovis
vaccine
peripheral blood
bovine
Prior art date
Application number
PCT/US1990/000106
Other languages
English (en)
Inventor
Lisle W. George
George M. Kagonyera
Original Assignee
The Regents Of The University Of California
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by The Regents Of The University Of California filed Critical The Regents Of The University Of California
Publication of WO1990007525A1 publication Critical patent/WO1990007525A1/fr

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • C07K14/21—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Pseudomonadaceae (F)
    • C07K14/212—Moraxellaceae, e.g. Acinetobacter, Moraxella, Oligella, Psychrobacter
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies

Definitions

  • This invention relates to a keratoconjunctivitis cytotoxin, to methods for extracting and purifying the cytotoxin and to veterinary vaccines comprising the cytotoxin.
  • IBK Infectious bovine keratoconjunctivitis
  • Moraxella bovis Moraxella sp. are aerobic, gram-negative rods appearing in pairs of short chains. They belong to the Neisseriaceae family and are oxidase (+) and catalase (-) .
  • IBK is highly contagious and can be transmitted rapidly throughout a herd. Acute to chronic inflammation of the eye occurs which impairs the animal's sight. This disease causes great financial loss in the cattle industry because it is a debilitating disease which affects cattle of all ages and breeds. *
  • Moraxella bovis is considered to be the principal infectious agent associated with IBK.
  • the earliest described ocular changes after inoculation of M ⁇ . bovis into the eyes of cattle include necrosis and sloughing of the corneal epithelium, and bacteria lying in pit-like defects on the surface of the epithelial cells.
  • the margins of these pit ⁇ like defects conform closely to the shape of the associated bacterial cell, indicating that epithelial cell cytotoxicity may constitute an important pathogenic mechanism in the early stages of IBK.
  • Macroscopically visible corneal ulcers may occur by 4 to 8 days after infection. At that time, the corneal stroma underlying the ulcerated area is infiltrated by bacteria and neutrophils. Many of the neutrophils and epithelial cells in the ulcerated areas appear necrotic.
  • the toxin disclosed herein is useful for a preventative vaccine.
  • This invention relates to a cytotoxin substantially free of intact microbial cells from Moraxella bovis which is toxic to bovine peripheral blood neutrophils and which lacks hemolytic activity.
  • This cytotoxin is useful as a vaccine for protection of animals from infectious bovine keratoconjunctivitis.
  • the cytotoxin is further characterized in that it is not capable of hydrolyzing casein, it is net negatively charged at a pH of 7.4 and by its ability to elute off of a DEAE exchange column with a 3ml gel bed under a salt gradient at a 0.2M t about a 0.3M salt concentration. Further, the cytotoxic activity to bovine peripheral blood neutrophils is sensitiv to zinc salt.
  • the cytotoxin is prepared by clarifying and purifying culture filtrates of Moraxella bovis.
  • ________ bovis which is cytotoxic to bovine peripheral blood neutrophils and which can be used as a vaccine against keratoconjunctivitis. Methods for purifying the cytotoxin are also disclosed. Extraction and Purification of the Cytotoxin
  • Moraxella bovis strains useful in preparing the cytotoxin of the invention can be isolated from clinical cases of IBK or can be obtained from public sources.
  • Publi sources include, for example, the American Type Culture Collection in Rockville, Maryland, U.S.A. (ATCC) where M. bovis strains are deposited under accession numbers 10900, 25576, 17947 and 17948. The most preferred strain is 5 " Tifto 1 strain, deposited with the ATCC on January 10, 1989 havin accession number 53854. The bacteria will grow on mos common bacterial culture media.
  • M. bovis grown on McCoy's media (Grand Island Biologicals, Grand Island, New York or Flow Labs, Inglewood, California) supplemented with about 3% bovine serum albumin (BSA) and about 3% fetal calf serum yields filtrates with high levels of cytotoxic activity.
  • BSA bovine serum albumin
  • a minimal maxim be used which contains fewer exogenous proteins.
  • Dialyzed GC media is a preferred minimal medium which yields good quantities of purified cytotoxin.
  • Dialyzed GC media contains all components of GC media which is commercially available except that high molecular weight proteose peptone is substituted with proteose peptone which has been solubilized in water and dialyzed to remove high M peptones, typically those greater than about 14,000M .
  • One liter of dialyzed GC media contains dialysate from 15g of powdered proteose peptone.
  • Cytotoxin production is best if the bacteria is grown on agar plates rather than in liquid shake cultures.
  • Agar plates or shake cultures are preferably incubated at a temperature of 37°C for about 18 to about 36 hours.
  • the bacteria is harvested from the plates by washing the agar surface with an inorganic buffer having a molarity of about 0.01 to 0.05 and a pH of 7.4.
  • the preferred buffer is Hepes and 200mM CaCl 2 .
  • the buffer is added to the agar plates in a ratio of about 5ml:75cm 2 to about 15ml:75cm 2 of agar surface area.
  • the culture filtrates are clarified by removing the solid materials by differential centrifugation, ultrafiltration or the like. Typically, the materials are centrifuged and the supernatant is then further purified by passing through a polycarbonate filter, preferably of about 0.22 ⁇ size.
  • the crude filtrate is next precipitated by using standard techniques known in the field, such as the use of cold acetone or salts. Concentrations and other conditions vary with the salt chosen. Standard experimental strategies can be used to optimize selective precipitation for each salt. Selective precipitation using solutes is preferred. The solute most preferred is 70% ammonium sulfate.
  • the filtrate may be centrifuged to collect the precipitate. Th precipitate is then collected and redissolved in a suitable buffer as described above.
  • Th first protein peak is collected and diluted at about 1:2 to about 1:10, most preferably about 1:10, in a buffer solution, as described above.
  • the preparation is then further purified using standard column chromatographic techniques which may include, for example, ion exchange chromatography, molecula filtration, electrophoresis, or isoelectric focusing.
  • Standard columns generally recognized for use in purifying proteins are contemplated in purifying the cytotoxin of the present invention and many alternative columns are commercially available and known to those skilled in the art.
  • the filtrate buffer solution is then placed on an anion exchange column such as those produced by Sigma Chemical Co., St. Louis, Missouri.
  • the anion exchange column is preferably a DEAE-Sephadex, DEAE-Sephacryl or DEA cellulose column having about a 1ml to 10ml gel bed. A DEA column with about a 3ml gel bed is most preferred.
  • the cytotoxin is eluted from the column through the use of a salt gradient.
  • the cytotoxin solution is bound to the column, the column is then washed with buffer and then a salt gradient is passed through the column.
  • the sal gradient may be of any range such that the range encompasse at least a 0.0M to 0.3M solution of a salt such as NaCl.
  • the purified cytotoxin elutes off of a DEAE exchange column under a salt gradient at about 0.2M to about 0.3M NaCl concentration.
  • eluate fraction may be individually tested to determine which fractions are cytotoxic to bovine peripheral blood neutrophils and which lack hemolytic activity. Characterization of the Cytotoxin
  • the M ⁇ . bovis cytotoxin of this invention is substantially free of intact microbial cells which means that it has been derived from ML. bovis cultures and filtered or otherwise purified so that it is about preferably 95% to 100%, most preferably 100%, free of intact microbial cells.
  • the M-_ bovis cytotoxin is cytotoxic to bovine peripheral blood neutrophils. This toxicity is measured by observing the release of chromium by the peripheral blood neutrophils upon subjection of the neutrophils to the cytotoxin pursuant to the methodology described by Miggiano et al. , Transpl. Proc.. 4_:231-237 (1972); Kurtz et al., Transfusion. .19.:398-403 (1979); and Carlson et al., Proc. Soc. Exp. Biol. Med. , 142:853-856 (1973).
  • the preferred method is set forth in Example II below.
  • the Mi. bovis cytotoxin is further characterized b its lack of hemolytic activity. Hemolytic activity can be assayed by methods known in the art such as that described by Bradley et al., J. Investi . Derm. ⁇ 78..206-209 (1982). The preferred method is that given in Example II below.
  • the cytotoxin is not capable of hydrolyzing casein.
  • the phrase "not capable of hydrolyzing casein” means that the cytotoxin, when placed in contact with a mil substrate, does not hydrolyze the casein and create clear zones in the media. Casein activity assays are described and well-known in the art as, for example, in U.S. Patent No. 4,675,176, which is incorporated by reference herein.
  • the cytotoxin is further characterized in that it cytotoxicity to bovine peripheral blood neutrophils is inhibited by zinc salts, such as zinc sulfate. When the cytotoxin is placed into contact with a zinc salt at a fina concentration of 2mM its cytotoxicity to the neutrophils is inhibited.
  • the cytotoxin has a molecular weight of about 42K based on polyacryla ide gel electrophoresis (PAGE) .
  • PAGE polyacryla ide gel electrophoresis
  • the cytotoxicity of the cytotoxin to bovine peripheral blood neutrophils is not inhibited when mixed with PMSF (40 ⁇ g/lml phenylmethane sulfonyl fluoride), pepstatin (0.7 ⁇ g/ml), TLC (50 ⁇ g/mi, l-l-chloro-3- [4-tosyl amido] -7-amino-2- heptanone-HCl) , phosphora idon (50 ⁇ g/ml) or leupeptin (0.5 ⁇ g/mi) either singly or together as a cocktail, all obtained from Boehringer Mannheim, Indianapolis, Indiana.
  • PMSF 40 ⁇ g/lml phenylmethane sulfonyl fluoride
  • the toxin is heat labile, being rapidly inactivated by heating at 56°C for over 15 minutes. It is net negatively charged molecule at a pH of 7.4.
  • the cytotoxins described herein from Moraxella bovis further include those which are immunologically equivalent to the cytotoxins derived from M ⁇ . bovis culture filtrates. By immunologically equivalent, it is meant that the cytotoxin embraces those that are immunologically indistinct to the immune systems of cattle. Such cytotoxin would include proteins identical to the cytotoxins describe herein as well as those comprising minor modifications to the primary sequence of the protein, such as amino acid deletions, additions, substitutions or chemical modifications thereto (e.g., alkylation, reduction).
  • the cytotoxins can be obtained from the cell cultures of M.
  • bovis but can also be derived from chemical synthetic mean or recombinant genetic technology.
  • To determine immunological equivalence one can use any number of standard immunological assays. The most convenient assay would involve competitive immunoassays wherein equivalence could be determined when antibodies against naturally secreted cytotoxin bind in about the same manner to about the same extent as the cytotoxin being assayed for equivalence.
  • a vaccine prepared utilizing the purified cytotoxin or its immunogenic equivalents thereof can be pre ⁇ pared in vaccine dose form by well-known procedures.
  • the vaccine can be administered intramuscularly or subcutaneously.
  • the immunogen may be combined with a suitable carrier, for example, it may be administered in water, saline or buffered vehicles with or without various adjuvants or immunomodulating agents such as aluminum hydroxide, aluminum phosphate, aluminum potassium sulfate (alum) , beryllium sulfate, silica, kaolin, carbon, water-in- oil emulsions, oil-in-water emulsions, muramyl dipeptide, bacterial endotoxin, lipid X, Corvnebacterium parvum (Propi ⁇ nobacterium acnes) , Bordetella pertussis, poly- ribonucleotides, sodium alginate, lanolin, lysolecithin, vitamin A
  • Such adjuvants are available commercially from various sources, for example, Merck Adjuvant 65 (Merck and Company, Irfc. , Rahway, N.J.). Other suitable adjuvants are Freund's Incom plete Adjuvant and Complete Adjuvant (Difco Laboratories, Detroit, Michigan) .
  • the proportion of immunogen and adjuvant can be varied over a broad range so long as both are present in effective amounts.
  • aluminum hydroxide can be present in an amount of about 0.5% of the vaccine mixture (A1 2 0 3 basis).
  • the concentration of the cytotoxin can range from about l ⁇ g to about 2,000 ⁇ g per bovine host.
  • a preferable range is from about lOO ⁇ g to about l,000 ⁇ g per dose.
  • a suitable dose size is about 1-lOml, preferably about 1.0ml.
  • a dose for intramuscular injection for example, would comprise 1ml containing l.O g of immunogen in admixture with 0.5% aluminum hydroxide.
  • Comparable dose forms can also be prepared for parenteral administration to calves, but the amount of immunogen per dose will be smaller, for example, about l ⁇ g to about 500 ⁇ g per dose.
  • a regimen of between 1 and 4 doses can be used with the injections spaced out over a two- to six-week period.
  • a two-dose regimen is used.
  • the second dose of the vaccine then should be administered some weeks after the first dose, for example, about two to four weeks later.
  • Animals that have been previously exposed to M. bovis or have received colostral antibodies from the mother may require booster injections.
  • a two-dose regimen is considered preferable for the most effective immunization o the calves.
  • Semiannual revaccination is recommended for breeding animals. Calves may be vaccinated at about 1-3 months after birth, again at four to six months, and yearly or preferably semi-annually thereafter.
  • the vaccine may also be combined with other vac ⁇ cines for other diseases to produce multivalent vaccines. It may also be combined with other medicaments, for example antibiotics.
  • a pharmaceutically effective amount of the vaccine can be employed with a pharmaceutically acceptable carrier or diluent understood to be useful for the vaccination of animals such as swine, cattle, sheep, goats, and other mammals.
  • vaccines may be prepared according to meth- ods well-known to those skilled in the art as set forth, fo example, in I. Tizard, "An Introduction to Veterinary Immu ⁇ nology,” 2nd Ed (1982), which is incorporated herein by ref erence.
  • the vaccine or the cytotoxin may be stored in lyophilized form. Though cytotoxic activity may be lost, the antigenic properties of the cytotoxin are maintained.
  • M_-_ bovis The hemolytic isolate of M_-_ bovis for these experiments was obtained from a cow with IBK in Georgia (Tifton 1) .
  • the bacterial isolate was maintained as lyophilized stock cultures. This isolate was positively identified as M. bovis using biochemical and morphologic criteria described in Smith and George, Am. J. Vet. Res. , 45:804-807 (1985) which is incorporated by reference herein.
  • the sacs were sealed at both ends and washed thoroughly in distilled water. The sacs were then placed in a graduated cylinder with parafilm to seal the top. The cylinders were then placed in the refrigerator. Dialysis was allowed to occur for 24-36 hours in the cold.
  • Hepes buffer (0.02M Hepes, lOOmg/l of calcium chloride) was pipetted into each of five, 15-ml screwcap pyrex tubes. Caps were screwed on each tube.
  • Isovitalex bottles were obtained from Becton-Dickinson Microbiology Systems. These bottles contain ingredients important for growing Gonorrhea coccus. thus the reason for the name GC agar. Typically, these bottles contain the following ingredients: vitamin B 12 , L-glutamine, guanine HC1, PABA, cocarboxylase adenine, diphosphoparadine nucleotide, oxidized (coenzyme 1) , ferric nitrate, thiamine, L-cysteine HC1, and L-cystine.
  • vitamins B 12 L-glutamine
  • guanine HC1 guanine HC1
  • PABA cocarboxylase adenine
  • diphosphoparadine nucleotide diphosphoparadine nucleotide
  • oxidized (coenzyme 1) ferric nitrate
  • thiamine thiamine
  • L-cysteine HC1 L-cystine
  • Isovi ⁇ tale TM li•qui «d was combined with the lyophilized powder per the instructions. After the powder was dissolved, the fluid was aspirated. 7. The agar, cooled to 56°C, was removed from the waterbath. The entire contents of the IsovitalexTM bottle was added to the erlenmeyer flask. About 300m of agar were poured into each petri plate. The agar was allowed to harden before inoculation. 8. 2-10 "fried egg” type of M bovis colonies were inoculated into the sterile Hepes solution. A 5cc pipette was used to swish the fluid back and forth one or two times.
  • the agar surface of each plate was flushed with 10ml of Hepes CaCl 2 (0.02M, lOOmg/1, pH 7.4) per 75cm 2 of the agar surface area.
  • a sterile glass hockey stick was used to remove the bacterial growth.
  • the harvested culture was pipetted into a 50-ml centrifuge tube. The tubes were centrifuged for 15 minutes at 3,000rpm. The supernatant was removed and filter-sterilized two times through a 0.22- micron polycarbonate filter (47mm diameter, Nucelopore, Inc., Pleasanton, California).
  • the sterile filtrate then was diluted 1:2 in the low ionic strength Hepes buffer, as described above, and 40ml was pumped onto a 3ml gel bed DEAE column (DEAE Spectra gel M, Spectrum Medical Industries Inc. , Los Angeles,
  • Bovine peripheral blood neutrophils were isolated by modification of methods described by Carlson et al., Proc. Soc. Exp. Biol. Med.. 142:853-856 (1973) and Chambers
  • Heparinized blood 40ml was collected from the jugular vein of either of 2 normal adult donors. After collection, 10ml aliquots of blood were centrifuged (4°C, 500xg) for 15 minutes. After centrifugation, the plasma was aspirated and
  • PBSS phosphate buffered saline solution
  • the suspended cells then were layered over 4.0ml of lymphocyte separation media (density 1.077 to l.080g/ml, Litton Bionetics Inc., Kensington, Maryland) and the mixture was centrifuged for 25 minutes (4°C, 500xg) .
  • lymphocyte separation media density 1.077 to l.080g/ml, Litton Bionetics Inc., Kensington, Maryland
  • the cells were counted using a hemocyt ⁇ meter and the viability was determined by staining with 0.1% eosin dye in phosphate buffered saline. After washing, the cells were resuspended and adjusted to a final concentration of 2xl0 7 neutrophils/ml by the addition of
  • the yield from each 40ml blood sample was approximately 5xl0 7 to 7xl0 7 cells, and was composed of 80% to 90% neutrophils. More than 90% of the cells were viable.
  • Isotope labeling of the neutrophils was performed according to Kurtz et al., Transfusion, l :398-403 (1979).
  • the purified neutrophil suspensions were incubated with 51 Cr labeled Na 2 Cr0 4 (100 ⁇ C/10 7 neutrophils, 37°C, 1 hour, ICN Inc. , Irvine, California) .
  • the cells were rotated (Roto-Torque, Cole Palmer, Chicago, ⁇ L) .
  • the cells were washed three times in cold PBSS and were finally resuspended in McCoy's to a final concentration of 1 x 10 6 neutrophils/ml before the radioactivity in 3 aliquots (0.5ml) of each cell suspension was counted (Autogamma Scintillation Spectrometer, Hewlett Packard, Palo Alto, California) .
  • the cytotoxicity assays were performed by incubating 0.5ml of the labeled neutrophils at 37°C with 0.5ml of the preparation to be assayed (crude l_ _ bovis filtrates or partially purified fractions) for 30 minutes with gentle rotation.
  • % Release cpm sample - cpm media controls x 2 x 100 (cpm neutrophils - cpm media controls)
  • McCoy's media 25mM Hepes buffer, Gibco Inc., Grand Island, 15 New York
  • Indicator substrate was 100ml phosphate buffer (0.6M, pH 6.0), 0.017ml of 30%H 2 O 2 , and 1.67mg/ml dianisidine (0.0167 g/lOOml buffer).
  • Protein content was measured using the commercially available Lowry's test kit (Sigma Chemical Co. St. Louis, Missouri) .
  • Table I sets forth the relative hemolytic and cytotoxic activity of fractions expressed in percent hemolysis or cytotoxicity.
  • O.D. represents the optical density of the fraction at a wavelength of 280nm. The measurement is linearly related to the protein concentration of the fraction.

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Medicinal Chemistry (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)

Abstract

Une cytotoxine tirée de la Moraxella bovis est essentiellement dépourvue de cellules microbiennes intactes, est toxique pour les neutrophiles sanguins périphériques de bovins et est dépourvue d'activité hémolytique. Cette cytotoxine est utile comme vaccin pour protéger des animaux de la kératonconjonctivite infectieuse bovine. La cytotoxine se caractérise en outre en ce qu'elle est incapable d'hydrolyser la caséine, qu'elle a une charge négative nette à un pH de 7,4 et qu'elle peut être éluée dans une colonne d'échange DEAE avec un lit colloïdal de 3ml au moyen d'un gradient salin ayant une concentration saline comprise entre 0,2M et 0,3M. En outre, l'activité cytotoxique sur les neutrophiles sanguins périphériques de bovins est sensible aux sels de zinc. Typiquement, on prépare la cytotoxine par clarification et par purification de filtrats de culture de Moraxella bovis.
PCT/US1990/000106 1989-01-06 1990-01-05 Cytotoxine de la keratoconjonctivite et ses procedes d'utilisation et de production WO1990007525A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US29423989A 1989-01-06 1989-01-06
US294,239 1989-01-06

Publications (1)

Publication Number Publication Date
WO1990007525A1 true WO1990007525A1 (fr) 1990-07-12

Family

ID=23132503

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US1990/000106 WO1990007525A1 (fr) 1989-01-06 1990-01-05 Cytotoxine de la keratoconjonctivite et ses procedes d'utilisation et de production

Country Status (2)

Country Link
AU (1) AU5088690A (fr)
WO (1) WO1990007525A1 (fr)

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996012502A1 (fr) * 1994-10-25 1996-05-02 Kansas State University Research Foundation Procede de culture de m.bovis dans un milieu a basse teneur en fer et preparation d'un vaccin contre la keratoconjonctivite infectieuse bovine a partir de ce procede
WO2001016172A1 (fr) * 1999-08-31 2001-03-08 Commonwealth Scientific And Industrial Research Organisation Antigenes de vaccin provenant de moraxella
WO2002102408A1 (fr) * 2001-06-19 2002-12-27 The Regents Of The University Of California Cytotoxine de moraxella bovis, gene de cytotoxine, anticorps et vaccins pour la prevention et le traitement d'infections dues a moraxella bovis
US9668677B2 (en) 2004-07-13 2017-06-06 Dexcom, Inc. Analyte sensor
US10524703B2 (en) 2004-07-13 2020-01-07 Dexcom, Inc. Transcutaneous analyte sensor
US10610135B2 (en) 2005-03-10 2020-04-07 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration

Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1986006635A1 (fr) * 1985-05-15 1986-11-20 Biotechnology Australia Pty. Ltd. Vaccins oraux

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1986006635A1 (fr) * 1985-05-15 1986-11-20 Biotechnology Australia Pty. Ltd. Vaccins oraux

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
AMERICAN JOURNAL OF VETERINARY RESEARCH, Volume 36(3), issued March 1975, HUGHES et al.: "Experimentally Induced Infectious Bovine Keratoconjuctivitis; Relationship of Vaccination Schedule to Protection Against Exposure with Homologous Moraella bouis Culture", pages 263-265. See the Summary and the last paragraph. *
AMERICAN JOURNAL OF VETERINARY RESEARCH, Volume 50(1), issued January 1989, KAGONYERA et al.: "Effects of Moraxella bovis and Culture filtrates on 51 Cr-labeled Bovine Neutrophils", pages 18-21. See the Discussion pages 20-21. *
AMERICAN JOURNAL OF VETERINARY RESEARCH, Volume 51(2), issued February 1990, HOIEN-DALEN et al.: "Comparative Characterization of the Leukocidic and Hemolytic Activity of Moraxella bocis", pages 191-196. See entire document. *
CANADIAN JOURNAL OF COMPARATIVE MEDICINE, Volume 37, issued January 1973, PUGH et al.: "The Pathophysiological Effects of Moraxella bovis Toxins on Cattle, Mice, an Guinea Pigs", pages 70-78. See the Abstract and pages 76-77. *
CONFERENCE OF RESEARCH WORKS IN ANIMAL DISEASE, Abstract Number 49, Published 1988, HOIEN-DALEN et al.: "Partial Characterization of a Moraxella bovis Leukocidin and Comparison to the M. bovis Hemolysin", page 9. See the entire abstract. *

Cited By (41)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1996012502A1 (fr) * 1994-10-25 1996-05-02 Kansas State University Research Foundation Procede de culture de m.bovis dans un milieu a basse teneur en fer et preparation d'un vaccin contre la keratoconjonctivite infectieuse bovine a partir de ce procede
WO2001016172A1 (fr) * 1999-08-31 2001-03-08 Commonwealth Scientific And Industrial Research Organisation Antigenes de vaccin provenant de moraxella
US7018639B1 (en) 1999-08-31 2006-03-28 Commonwealth Scientific And Industrial Research Organisation Vaccine antigens of Moraxella
WO2002102408A1 (fr) * 2001-06-19 2002-12-27 The Regents Of The University Of California Cytotoxine de moraxella bovis, gene de cytotoxine, anticorps et vaccins pour la prevention et le traitement d'infections dues a moraxella bovis
US7220421B2 (en) * 2001-06-19 2007-05-22 The Regents Of The University Of California Moraxella bovis cytotoxin, cytotoxin gene, antibodies and vaccines for prevention and treatment of Moraxella bovis infections
US10709363B2 (en) 2004-07-13 2020-07-14 Dexcom, Inc. Analyte sensor
US10993642B2 (en) 2004-07-13 2021-05-04 Dexcom, Inc. Analyte sensor
US11883164B2 (en) 2004-07-13 2024-01-30 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US11064917B2 (en) 2004-07-13 2021-07-20 Dexcom, Inc. Analyte sensor
US11045120B2 (en) 2004-07-13 2021-06-29 Dexcom, Inc. Analyte sensor
US11026605B1 (en) 2004-07-13 2021-06-08 Dexcom, Inc. Analyte sensor
US10993641B2 (en) 2004-07-13 2021-05-04 Dexcom, Inc. Analyte sensor
US10918314B2 (en) 2004-07-13 2021-02-16 Dexcom, Inc. Analyte sensor
US10709362B2 (en) 2004-07-13 2020-07-14 Dexcom, Inc. Analyte sensor
US10524703B2 (en) 2004-07-13 2020-01-07 Dexcom, Inc. Transcutaneous analyte sensor
US10722152B2 (en) 2004-07-13 2020-07-28 Dexcom, Inc. Analyte sensor
US10980452B2 (en) 2004-07-13 2021-04-20 Dexcom, Inc. Analyte sensor
US10799159B2 (en) 2004-07-13 2020-10-13 Dexcom, Inc. Analyte sensor
US10799158B2 (en) 2004-07-13 2020-10-13 Dexcom, Inc. Analyte sensor
US10813576B2 (en) 2004-07-13 2020-10-27 Dexcom, Inc. Analyte sensor
US10932700B2 (en) 2004-07-13 2021-03-02 Dexcom, Inc. Analyte sensor
US10827956B2 (en) 2004-07-13 2020-11-10 Dexcom, Inc. Analyte sensor
US9668677B2 (en) 2004-07-13 2017-06-06 Dexcom, Inc. Analyte sensor
US10918313B2 (en) 2004-07-13 2021-02-16 Dexcom, Inc. Analyte sensor
US10918315B2 (en) 2004-07-13 2021-02-16 Dexcom, Inc. Analyte sensor
US10925524B2 (en) 2005-03-10 2021-02-23 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10743801B2 (en) 2005-03-10 2020-08-18 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10918316B2 (en) 2005-03-10 2021-02-16 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10918317B2 (en) 2005-03-10 2021-02-16 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10898114B2 (en) 2005-03-10 2021-01-26 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10856787B2 (en) 2005-03-10 2020-12-08 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10610135B2 (en) 2005-03-10 2020-04-07 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10918318B2 (en) 2005-03-10 2021-02-16 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10716498B2 (en) 2005-03-10 2020-07-21 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10709364B2 (en) 2005-03-10 2020-07-14 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US11000213B2 (en) 2005-03-10 2021-05-11 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10617336B2 (en) 2005-03-10 2020-04-14 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10610136B2 (en) 2005-03-10 2020-04-07 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US11051726B2 (en) 2005-03-10 2021-07-06 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10610137B2 (en) 2005-03-10 2020-04-07 Dexcom, Inc. System and methods for processing analyte sensor data for sensor calibration
US10813577B2 (en) 2005-06-21 2020-10-27 Dexcom, Inc. Analyte sensor

Also Published As

Publication number Publication date
AU5088690A (en) 1990-08-01

Similar Documents

Publication Publication Date Title
Osserman et al. Serum and urinary lysozyme (muramidase) in monocytic and monomyelocytic leukemia
Lancefield et al. The properties of T antigens extracted from group A hemolytic streptococci
Roberts The relationship between group A and group C meningococcal polysaccharides and serum opsonins in man
Valle et al. Immune specific production of interferon by human T cells in combined macrophage-lymphocyte cultures in response to herpes simplex antigen
Rottini et al. Identification and partial characterization of a cytolytic toxin produced by Gardnerella vaginalis
Gaines et al. Prevalence of antibody to Pseudomonas in normal human sera
Taranta et al. Dissociation of hemolytic and lymphocyte-transforming activities of streptolysin S preparations
Goodman et al. Endotoxin protein is a mitogen and polyclonal activator of human B lymphocytes.
Beachey et al. Toxic effects of streptococcal M protein on platelets and polymorphonuclear leukocytes in human blood
Cunningham et al. Suppression of antibody response by group A streptococcal pyrogenic exotoxin and characterization of the cells involved
Wyler et al. Response of sensitized and unsensitized human lymphocyte subpopulations to Plasmodium falciparum antigens
Kantor et al. Preparation and antigenicity of M protein released from group A, type 1 streptococcal cell walls by phage-associated lysin
Bailey et al. Antigenic properties of pleuropneumonia-like organisms from tissue cell cultures and the human genital area
Halbert THE USE OF PRECIPITIN ANALYSIS IN AGAR FOR THE STUDY OF HUMAN STREPTOCOCCAL INFECTIONS: III. THE PURIFICATION OF SOME OF THE ANTIGENS DETECTED BY THESE METHODS
Ogburn et al. Extracellular antigens in steady-state cultures of the hemolytic streptococcus: production of proteinase at low pH
Janney et al. Cold hemagglutinin cross-reactivity with Mycoplasma pneumoniae
Mendes et al. T and B lymphocytes in patients with lepromatous leprosy
WO1990007525A1 (fr) Cytotoxine de la keratoconjonctivite et ses procedes d'utilisation et de production
Pierce Electrophoretic and Immunological Studies on Sera from Calves from Birth to Weaning II. Electrophoretic and Serological Studies with Special Reference to the Normal and Induced Agglutinins to Trichomonas Foetus
Keiser et al. “Nonspecific” stimulation of lymphocyte transformation by cellular fractions and acid extracts of group A streptococci
Heilman et al. TISSUE CULTURE STUDIES ON BACTERIAL ALLERGY IN EXPERIMENTAL BRUCELLOSIS: I. The Effect of Brucella suis Whole Antigen on Cultures of Spleen from Normal and Brucella-Infected Guinea Pigs
Barta et al. Inhibition of lymphocyte blastogenesis by whey
Rantz et al. Antibacterial precipitating antibodies in group A hemolytic streptococcus sore throat
Rudner et al. Systemic and topical protection studies using Pseudomonas aeruginosa flagella in an ocular model of infection
Kaplan Rheumatic fever, rheumatic heart disease, and the streptococcal connection: the role of streptococcal antigens cross-reactive with heart tissue

Legal Events

Date Code Title Description
AK Designated states

Kind code of ref document: A1

Designated state(s): AT AU BB BG BR CA CH DE DK FI GB HU JP KP KR LK LU MC MG MW NL NO RO SD SE SU

AL Designated countries for regional patents

Kind code of ref document: A1

Designated state(s): AT BE BF BJ CF CG CH CM DE DK ES FR GA GB IT LU ML MR NL SE SN TD TG

REG Reference to national code

Ref country code: DE

Ref legal event code: 8642

NENP Non-entry into the national phase

Ref country code: CA

点击 这是indexloc提供的php浏览器服务，不要输入任何密码和下载