Journal article
High-throughput identification of T-lymphocyte antigens from Anaplasma marginale expressed using in vitro transcription and translation
Journal of immunological methods, Vol.332(1), pp.129-141
2008
Handle:
https://hdl.handle.net/2376/105540
PMID: 18243240
Abstract
The ability to rapidly screen a complex pathogen proteome for proteins that elicit recall T-lymphocyte responses from immune individuals would accelerate vaccine development. An outer membrane fraction of the rickettsial pathogen
Anaplasma marginale induces protective immunity against infection and disease in cattle. We have used this immunization model to evaluate high-throughput screening of proteins expressed by in vitro transcription and translation (IVTT) for recognition by memory CD4
+ T-lymphocytes. Fifty selected vaccine candidate antigens identified from the
A. marginale genome were expressed from transcriptionally active PCR products using an
Escherichia coli-based IVTT system, and bead-affinity purified using antibodies to His and FLAG epitope tags. IVTT-expressed bead-bound antigens were processed and presented by antigen presenting cells to T-lymphocytes from outer membrane immunized animals and evaluated for immunogenicity in proliferation assays. Antigens that consistently stimulated responses were known T-cell antigens major surface protein (MSP)2, MSP3, VirB9, and VirB10 and newly identified T-cell antigens outer membrane protein (OMP)4, OMP9, elongation factor-Tu, Ana29, and OMA87. Specific T-cell stimulation was achieved even at low antigen concentration, and was highly sensitive when compared with unbound IVTT reaction products. This method allows rapid expression and identification of T-lymphocyte antigens for any pathogen for which the genome sequence is available.
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Details
- Title
- High-throughput identification of T-lymphocyte antigens from Anaplasma marginale expressed using in vitro transcription and translation
- Creators
- Job E Lopez - Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, United StatesPaul A Beare - Coxiella Pathogenesis Section, Laboratory of Intracellular Parasites, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, MT 59840, United StatesRobert A Heinzen - Coxiella Pathogenesis Section, Laboratory of Intracellular Parasites, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, MT 59840, United StatesJunzo Norimine - Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, United StatesKevin K Lahmers - Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, United StatesGuy H Palmer - Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, United StatesWendy C Brown - Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, United States
- Publication Details
- Journal of immunological methods, Vol.332(1), pp.129-141
- Academic Unit
- Veterinary Microbiology and Pathology, Department of; Paul G. Allen School for Global Animal Health
- Publisher
- Elsevier B.V
- Identifiers
- 99900546662201842
- Language
- English
- Resource Type
- Journal article