| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 2 | St. Edward's |
A | Cocks | Taft | Donohoe | Niemi | 3:54.61 | Viera High School Invitational |
| 3 | Port St. Lucie |
A | Davis | Nelson | Wright | Bradley | DNS | Rockledge Raider Classic |
| 4 | Centennial |
A | Dimarco | Barber | Kirkwood | Johnson | 3:22.85 | Cobra Relays 2025 |
| 5 | Fort Pierce Central |
A | Ransfer | Hughes | Filsaine | Pascoe | 3:36.21 | Cobra Relays 2025 |
| 6 | Martin County |
A | Allison | Brown | Treadwell | Zelaya | DNS | Bulldog Invitational 2025 |