| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Blake |
A | NT | |
| 2 | Sickles |
A | 43.26 | 43.26 WC American Divisonals |
| 3 | Lennard |
A | 42.82 | 42.82 2023 Manuel-Griffin-Fraundorfer Relays |
| 4 | Armwood |
A | 41.16 | FHSAA Class 4A Track & Field State Championships |
| 5 | Hillsborough |
A | 46.00 | |
| 6 | Plant City |
A | 53.00 | |
| 7 | Alonso |
A | NT |
| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Middleton |
A | 44.58 | FHSAA 3A Region 3 |
| 2 | Strawberry Crest |
A | 43.98 | 43.98 WC National Division Meet |
| 3 | Sumner |
A | 50.00 | |
| 4 | Brandon |
A | DQ | |
| 5 | Robinson |
A | 43.50 | |
| 6 | Leto |
A | NT | |
| 7 | Newsome |
A | NT |
| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 2 | Riverview |
A | DNS | |
| 3 | Wiregrass Ranch |
A | DNS | East Lake Early Bird Invitational |
| 4 | Bloomingdale |
A | DNS | FHSAA Class 4A Track & Field State Championships |
| 5 | Tampa Bay Tech |
A | DNS | |
| 6 | Freedom (Tampa) |
A | DNS | |
| 7 | Spoto |
A | DNS |