| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 1 | Calvary Christian (PtStL) | A | Brown | Andino | Clark | Gonzalez | NT | |
| 2 | Tradition Prep | A | Hamilton | Chavers | Falcon | Parrel | NT | |
| 3 | St. Edward's |
A | Elliott | Gayle | Jensen | Perez | 55.43 | Viera High School Invitational |
| 4 | Archbishop McCarthy |
A | Hughes | Cabanban | Boursiquot | Sebelen | DQ | Pine Crest Panther Relays |
| 5 | Martin County |
A | Edwards | Perez | Garcia | Fernandes | 53.99 | Dr Garcia Invite #1 2025 |
| 6 | Treasure Coast | A | Charles | Braziel | Labbe | Smith | NT | |
| 7 | Jupiter |
A | Palomba | Granda | Stewart | Monterroso | NT |