A new adenovirus from sheep.
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Adenoviridae↗
Biomedical subjects
J M Sharp
Publications and source records attributed to J M Sharp.
Strategies for enhancing monoclonal antibody accumulation in plant cell and organ cultures.
Various strategies aimed at improving IgG(1) antibody accumulation in transgenic tobacco cell and organ cultures were tested. The form of tissue had a significant effect on antibody levels; shooty teratomas were less productive than hairy roots or suspended cells. Although there were several disadvantages associated with hairy roots compared with suspensions, such as slower growth, slower antibody production, and formation of a greater number of antibody fragments, the roots exhibited superior long-term culture stability. Antibody accumulation in hairy root cultures was improved by increasing the dissolved oxygen tension to 150% air saturation, indicating the need for effective oxygen transfer in root reactors used for antibody production. Preventing N-linked glycosylation using tunicamycin or inhibition of subsequent glycan processing by castanospermine reduced antibody accumulation in the biomass and/or medium in cell suspensions. Loss of antibody from the cultures after its secretion and release into the medium was identified as a major problem. This effect was minimized by inhibiting protein transport in the secretory pathway using Brefeldin A, resulting in antibody accumulation levels up to 2.7 times those in untreated cells. Strategies for protecting secreted antibody, such as addition of poly(vinylpyrrolidone) and periodic harvesting from the medium using hydroxyapatite resin, also increased antibody titers. The mechanisms responsible for the disappearance of antibody from plant culture media were not clearly identified; degradation by proteases and conformational modification of the antibody, such as formation of aggregates, provided an explanation for some but not all the phenomena observed. This work demonstrates that the manipulation and control of culture conditions and metabolic processes in plant tissue cultures can be used to improve the production of foreign proteins. However, loss of secreted antibody from plant culture medium is a significant problem that may limit the feasibility of using product recovery from the medium to reduce downstream processing costs relative to agricultural systems.
Animals↗
Association of jaagsiekte sheep retrovirus with pulmonary carcinoma in Sardinian moufflon (Ovis musimon).
Bronchiolo-alveolar carcinoma has been described in man and in several animal species, including cattle, dogs, opossums, goats and sheep. In sheep, a bronchiolo-alveolar carcinoma, known as ovine pulmonary carcinoma (OPC), is caused by jaagsiekte sheep retrovirus (JSRV), an exogenous type D retrovirus. In the mid-1980s, a severe outbreak of a disease resembling OPC was described in captive Sardinian moufflon (Ovis musimon). In the present study, the use of polymerase chain reaction (PCR) amplification of nucleic acids extracted from archival material established that JSRV was associated with OPC in affected moufflon. JSRV was detected in the lungs and mediastinal lymph nodes. Immunohistochemical and in-situ PCR demonstrated that in the lungs, JSRV proviral DNA was localized in transformed and untransformed type II pneumocytes and in the alveolar macrophages. In the mediastinal lymph nodes, JSRV DNA was mainly located in the cortical follicles and paracortex. These data suggest that JSRV is the cause of OPC in Sardinian moufflon, as it is in Sardinian sheep.
Animals↗
Antibodies to ovine adenoviruses in sheep in Scotland.
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Adenoviridae↗
Detection of PCR products from Mycobacterium avium subspecies Paratuberculosis using oligonucleotides containing multiple 2,4-dinitrophenyl reporter groups.
A pool of five oligonucleotides has been used to detect the pathogenic organism Mycobacterium avium subspecies paratuberculosis in PCR-amplified DNA from ruminants. The oligonucleotides were labelled at the 5'-end with three dinitrophenyl reporter groups and hybridised to the target DNA, which was fixed to a nylon membrane by ultraviolet irradiation. Colourimetric detection of the PCR product was carried out using an anti-DNP antibody conjugated to horseradish peroxidase or to alkaline phosphatase. Detection with alkaline phosphatase was more sensitive than with horseradish peroxidase but, in both cases, the PCR product could be easily detected. The DNP labelling system offers an economic and effective alternative to biotin, digoxigenin or fluorescein for the detection of PCR-amplified DNA.
Bacterial Typing Techniques↗