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US 9,872,896 B2Granted
Immortalized porcine alveolar macrophage
AssigneeIntervet Inc.
InventorsJaap Kool, Carla Christina Schrier
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Abstract
The present invention relates to immortalized porcine alveolar macrophages (PAMs), to cell cultures comprising such PAMs, to methods for the immortalization of PAMs, to methods of replicating PRRS virus on immortalized PAMs and to methods for the preparation of vaccines comprising PRRSV.
Claims11
A method of isolating progeny PRRSV from an immortalized porcine alveolar macrophage (PAM), wherein said PAM is susceptible to Porcine Respiratory and Reproductive Virus (PRRSV), expresses an SV40 T antigen, and does not comprise retroviral Long Terminal Repeat DNA capable of retroviral gene expression;
- wherein said method comprises the steps of:
- a) obtaining a cell-containing bronchoalveolar lavage sample from a porcine subject,
- b) separating a cellular component from said sample,
- c) transfecting said cellular component with a DNA molecule comprising transposons and comprising a gene encoding the SV40 T antigen under the control of a suitable promoter,
- d) selecting immortalized PAM cells that have been cultured for at least 5 cell cycles,
- e) further culturing the selected immortalized PAM cells,
- f) contacting the immortalized PAM cells with the PRRSV,
- g) allowing the PRRSV to replicate and form progeny PRRSV, and
- h) isolating the progeny PRRSV.
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Dependencies are inferred from the claim text.