Table 3.
T Cell Response to AAV7 Capsid in Peripheral Blood Mononuclear Cells and Liver-Derived Lymphocytes Detected by Either ex Vivo or Cultured Enzyme-Linked Immunospot Assaya
| |
|
T cell response to AAV capsid (SFU/106cells) |
||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| |
|
Ex vivo |
|
|
|
|
|
|||||
| |
|
PBMCs |
|
Cultured PBMCs |
||||||||
| Animal no. | Necropsy (day) | Pre | Day 16 | Day 23 | Day 30 | End | Liver | Pre | Day 16 | Day 23 | Day 30 | End |
| 24466 | 8 | (–) | NA | NA | NA | (–) | (–) | (–) | NA | NA | NA | ND |
| 24527 | 8 | (–) | NA | NA | NA | (–) | (–) | (–) | NA | NA | NA | ND |
| 24441 | 8 | (–) | NA | NA | NA | (–) | (–) | (–) | NA | NA | NA | ND |
| 24456 | 8 | (–) | NA | NA | NA | (–) | (–) | (–) | NA | NA | NA | ND |
| 24499 | 8 | (–) | NA | NA | NA | (–) | (–) | (–) | NA | NA | NA | ND |
| 24462 | 8 | (–) | NA | NA | NA | (–) | 671b | (–) | NA | NA | NA | ND |
| 24461 | 37 | (–) | (–) | (–) | (–) | (–) | 345c | (–) | (–) | (–) | (–) | (–) |
| 24479 | 37 | (–) | (–) | (–) | (–) | (–) | (–) | (–) | 125b | (–) | (–) | ND |
| 24481 | 37 | (–) | (–) | (–) | (–) | (–) | (–) | (–) | 135b | 454d | 570d | ND |
Abbreviations: AAV, adeno-associated virus: NA, not applicable; ND, not determined; PBMCs, peripheral blood mononuclear cells; SFU, spot-forming unit.
Isolation of PBMCs and liver-derived lymphocytes and ELISPOT assays are described in Materials and Methods. A peptide library specific for the full length of AAV7-VP1 was synthesized as 15-mers with 11-amino acid overlaps (Mimotopes, Clayton, Victoria, Australia). The entire lyophilized library, consisted of 146 peptides, was reconstituted with dimethyl sulfoxide and grouped into pools A (50 peptides). B (50 peptides), and C (46 peptides) for use in cell stimulation. Samples were harvested before injection (Pre), at necropsy (End), and, for the day 37 animals, on days 16, 23, and 30. Please see additional footnotes in Table 2.
AAV7 peptide pool C.
AAV7 peptide pool A.
AAV7 peptide pool A + B + C.