| Chapter title |
Dendritic Cell-Based ELISpot Assay for Assessing T-Cell IFN-γ Responses in Human Peripheral Blood Mononuclear Cells to Dengue Envelope Proteins
|
|---|---|
| Chapter number | 17 |
| Book title |
Handbook of ELISPOT
|
| Published in |
Methods in molecular biology, January 2018
|
| DOI | 10.1007/978-1-4939-8567-8_17 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-8566-1, 978-1-4939-8567-8
|
| Authors |
Peifang Sun, Monika Simmons, Sun, Peifang, Simmons, Monika |
| Abstract |
Dengue envelope (E) protein is a dominant antigen for vaccine development and E-based vaccines have shown partial or full protection against live-virus challenge in non-human primates. Generally, T cell responses can be investigated with peptides. However, hundreds of over-lapping peptides need to be synthesized to cover the whole sequence of a protein, which brings the cost up to a much higher level than purchasing a protein. We have developed an enzyme-linked immunospot (ELISpot) assay that uses intact E proteins instead of peptides for assessing IFN-gamma (IFN-γ) responses. The assay relies on professional antigen presenting cells, dendritic cells, to process and present the E proteins to stimulate T cells.Peripheral blood mononuclear cells (PBMCs) from dengue-exposed and naïve subjects were selected for the assay development. IFN-γ production ranged from 53 to 513 spot forming units (SFUs) and 0-45 SFUs per million PBMCs in dengue-exposed and naive subject groups, respectively. The assay allowed quantification of E-specific IFN-γ secreting memory T cells in subjects 9 years after exposure to a live-attenuated virus vaccine and live-virus challenge. Our results suggest that the dendritic cell-based IFN-γ assay is a useful tool for assessing immunological memory for clinical research. |
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Mendeley readers
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|---|---|---|
| Unknown | 14 | 100% |
Demographic breakdown
| Readers by professional status | Count | As % |
|---|---|---|
| Student > Bachelor | 3 | 21% |
| Researcher | 2 | 14% |
| Unspecified | 1 | 7% |
| Student > Doctoral Student | 1 | 7% |
| Student > Master | 1 | 7% |
| Other | 0 | 0% |
| Unknown | 6 | 43% |
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|---|---|---|
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| Nursing and Health Professions | 1 | 7% |
| Unspecified | 1 | 7% |
| Immunology and Microbiology | 1 | 7% |
| Other | 1 | 7% |
| Unknown | 5 | 36% |