| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 3 | Freedom (Orlando) |
A | 45.65 | Harmony Invitational |
| 4 | Boone |
A | NT | |
| 5 | Windermere Prep |
A | DNS |
| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 2 | Horizon |
A | 44.95 | Windermere Meet of Champions Classic 2024 |
| 3 | Winter Park |
A | 43.50 | |
| 4 | Apopka |
A | 42.73 | |
| 5 | Olympia |
A | 42.81 | Windermere Meet of Champions Classic 2024 |
| 6 | West Orange |
A | 43.67 | Windermere Meet of Champions Classic 2024 |
| 7 | West Oaks |
A | 45.19 | Olympia Titans Invitational |
| 8 | Foundation Academy |
A | NT |