| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Imagine School |
A | NT | |
| 2 | Babcock | A | NT | |
| 3 | Braden River |
A | 55.36 | |
| 4 | Hardee |
A | 54.01 | FHSAA 2A Region 3 |
| 5 | Bradenton Christian |
A | DNS | 54.16 Ascender Opener 2 |
| 6 | Out-of-Door |
A | 55.77 | 55.77 Sarasota County Track and Field Championship |
| 7 | Sarasota Military Academy |
A | NT |
| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Sarasota |
A | 52.69 | 52.69 IMG Academy Invitational |
| 2 | Lakewood Ranch |
A | 51.89 | FHSAA 4A Region 2 |
| 3 | Parrish |
A | DQ | |
| 4 | Southeast |
A | 49.04 | Pepsi Florida Relays |
| 5 | Manatee |
A | DNF | 49.45 Lakewood Ranch Invite 2024 |
| 6 | Cardinal Mooney |
A | 51.43 | 51.43 Cardinal Mooney Invitational |
| 7 | North Port |
A | DQ | |
| 8 | Palmetto |
A | 53.00 |