| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 2 | Sarasota |
A | 54.29 | 54.29 Sarasota Riverview - Ram Invite 2025 |
| 3 | Lemon Bay |
A | 52.58 | 52.58 Sarasota County Track and Field Championship |
| 4 | Imagine School |
A | DQ | Dunbar Tiger Classic Invite (Approved Qualifier) |
| 5 | Out-of-Door |
A | 56.00 |
| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Riverview (Sarasota) |
A | 50.15 | FHSAA 4A Region 3 |
| 2 | Cardinal Mooney |
A | 49.58 | 49.58 Larry Richardson Memorial Invitational |
| 3 | Charlotte |
A | 49.03 | FHSAA 3A Region 3 |
| 4 | North Port |
A | 49.10 | |
| 5 | Parrish |
A | 50.00 | |
| 6 | Venice |
A | 51.69 | Larry Richardson Memorial Invite |