Table 1.
Epitope mapping of anti-IN–scFvs
| Peptide # | Amino acid position | Amino acid sequence | 7 | 104 | 135 | ScFv# 142 | 144 | BM10 | 756 | IN anti serum 758 |
|---|---|---|---|---|---|---|---|---|---|---|
| 4324 | 1–5 | EQVDKLVSAGIRKVLFLDGI | − | − | − | − | − | − | + | − |
| 4325 | 6–15 | IRKVLFLDGIDKAQDEHEKY | − | − | − | − | − | − | + | − |
| 4326 | 6–25 | IRKVLFLDGIDKAQDEHEKY | − | − | − | − | − | − | + | − |
| 4327 | 16–35 | HSNWRAAIASDFNLPPVVAKE | − | − | − | − | − | − | − | − |
| 4328 | 26–45 | FNLPPVVAKEIVASCDKCQL | − | − | − | − | − | − | − | − |
| 4329 | 36–55 | IVASCDKCQLKGEAMHGQVD | − | − | − | − | − | − | − | − |
| 4330 | 46–65 | KGEAMHGQVDCSPGIWQLDC | − | − | − | − | − | − | − | − |
| 4331 | 56–75 | CSPGIWQLDCTHLEGKYILV | − | − | − | − | − | − | − | − |
| 4332 | 66–85 | THLEGKVILVAVHVASGYIE | − | − | − | − | − | − | − | − |
| 4333 | 76–95 | AVHVASGYIEAEVIPAETGQ | − | − | − | − | − | − | − | − |
| 4334 | 86–105 | AEVIPAETGQETAYFLLKLA | − | − | − | − | − | − | − | − |
| 4335 | 96–115 | ETAYFLLKLAGRWPYKTIHT | − | − | − | − | − | − | − | − |
| 4336 | 106–125 | GRWPVKTIHTDNGSNFTGAT | − | − | − | − | − | − | − | − |
| 4337 | 116–135 | DNGSNFTGATVRAACWWAGI | − | − | − | − | − | − | − | − |
| ||||||||||
| 4339 | 136–155 | KQEFGIPYNPQSQGVVESMN | − | − | − | − | − | − | − | − |
| 4340 | 146–165 | QSQGYYESMNKELKKIIGQY | − | − | − | − | − | − | − | − |
| 4341 | 156–175 | KELKKIIGOY’RDQAEHLKTA | − | − | − | − | − | − | − | − |
| 4342 | 166–185 | RDQAEHLKTAYQMAYFIHNF | − | − | − | − | − | − | − | − |
 ; | ||||||||||
| 4344 | 186–205 | KRKGGIGGYSAGERIVDIIA | − | − | − | − | − | − | − | − |
| 4345 | 196–215 | AGERIVDIIATDIOTKELQK | − | − | − | − | − | − | − | − |
 ; | ||||||||||
| 4348 | 226–245 | YYRDSRXPLWKGPAKLLWKG | − | − | − | − | − | − | − | − |
| 4349 | 236–255 | KGPAKLLWKGEGAVVIQDNS | − | − | − | − | − | − | − | − |
| 4350 | 246–265 | EGAVVIQDNSDIKVVPRRKA | − | − | − | − | − | − | − | − |
| 4351 | 256–275 | DIKVVPRRKAKIIRDYGKQM | − | − | − | − | − | − | − | − |
| 4352 | 266–285 | KIIRDYGKQMAGDDCVASRQ | − | − | − | − | − | − | − | + |
| 4353 | 276–288 | AGDDCVASRQDED | − | − | − | − | − | − | − | + |
To perform epitope mapping, purified anti-IN–scFv clones 7, 104, 135, 142, and 144 were tested for binding to a set of 30 overlapping synthetic peptides covering the entire HIV-1 IN protein sequence. CovaLink ELISA plates (Thermo Fisher Scientific) were coated with 5 μg of each peptide and incubated overnight at 4 °C. The plates were then blocked with BSA and incubated with purified antibody fragments. After washing the wells with low-ionic-strength buffer or high-ionic-strength buffer (see Materials and methods), HRP-conjugated anti-HA mAb was used for detection. The same binding pattern was observed in both washing conditions. The anti-LANA1 scFv (BM10) was used as a control antibody. Integrase antiserum #756 and #758 were used to control for quality of peptides and ELISA conditions. Black boxes highlight the IN peptides bound by scFvs. The (+) symbol indicates binding and the (−) symbol means no binding.