Cold-adapted equine influenza viruses
a technology for equine influenza and cold-adapted foals, which is applied in the field of experimentally generated cold-adapted equine influenza viruses, can solve the problems of undesirable side effects, minimal protection for horses, and current modalities that cannot be used in young foals, and achieve the effect of reducing nasal discharge and temperatur
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example 1
[0062] This example discloses the production and phenotypic characterization of several cold-adapted equine influenza viruses of the present invention.
[0063] A. Parental equine influenza virus, A / equine / Kentucky / 1 / 91 (H3N8) (obtained from Tom Chambers, the University of Kentucky, Lexington, Ky.) was subjected to cold-adaptation in a foreign host species, i.e., embryonated chicken eggs, in the following manner. Embryonated, 10 or 11-day old chicken eggs, available, for example, from Truslow Farms, Chestertown, Md. or from HyVac, Adel, Iowa, were inoculated with the parental equine influenza virus by injecting about 0.1 milliliter (ml) undiluted AF containing approximately 106 plaque forming units (pfu) of virus into the allantoic cavity through a small hole punched in the shell of the egg. The holes in the eggs were sealed with nail polish and the eggs were incubated in a humidified incubator set at the appropriate temperature for three days. Following incubation, the eggs were cand...
example 2
[0072] Therapeutic compositions of the present invention were produced as follows.
[0073] A. A large stock of EIV-P821 was propagated in eggs as follows. About 60 specific pathogen-free embryonated chicken eggs were candled and non-viable eggs were discarded. Stock virus was diluted to about 1.0×105 pfu / ml in sterile PBS. Virus was inoculated into the allantoic cavity of the eggs as described in Example 1A. After a 3-day incubation in a humidified chamber at a temperature of about 34° C., AF was harvested from the eggs according to the method described in Example 1A. The harvested AF was mixed with a stabilizer solution, for example A1 / A2 stabilizer, available from Diamond Animal Health, Des Moines, Iowa, at 25% V / V (stabilizer / AF). The harvested AF was batched in a centrifuge tube and was clarified by centrifugation for 10 minutes at 1000 rpm in an IEC Centra-7R refrigerated table top centrifuge fitted with a swinging bucket rotor. The clarified fluid was distributed into 1-ml cryo...
example 3
[0076] A therapeutic composition comprising cold-adapted equine influenza virus EIV-P821 was tested for safety and its ability to replicate in three horses showing detectable prior immunity to equine influenza virus as follows. EIV-P821, produced as described in Example 1A, was grown in eggs as described in Example 2A and was formulated into a therapeutic composition comprising 107 pfu EIV-P821 / 2 ml BSA-MEM solution as described in Example 2C.
[0077] Three ponies having prior detectable hemagglutination inhibition (HAI) titers to equine influenza virus were inoculated with a therapeutic composition comprising EIV-P821 by the following method. Each pony was given a 2-ml dose of EIV-P821, administered intranasally using a syringe fitted with a blunt cannula long enough to reach past the false nostril, 1 ml per nostril.
[0078] The ponies were observed for approximately 30 minutes immediately following and at approximately four hours after vaccination for immediate type allergic reactio...
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