| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 1 | Cornerstone Charter |
A | Gainex | Tibo | Hierbe | Janis | NT | |
| 2 | Innovation_Orlando | A | McNeill-Toro | Londoni | Batts | Turner | 53.25 | Lake Nona - Tammy Lafayette Relays 2025 |
| 3 | Evans |
A | Clempson | Mobley | Sullivan | Prophete Ross | 50.95 | Blue Devil Invitational (Regio |
| 4 | Dr. Phillips |
A | Castell | Smith | Martin | Kirkland | 48.83 | Charlie Harris Relay |
| 5 | Edgewater |
A | Pennicook | Colbert | Todd | Green | DNS | Tiger Invitational |
| 6 | Bishop Moore |
A | Suffran | Sullivan | Maddalena | Sweeney | 52.23 | ROYAL INVITATIONAL @ The First Academy 2025 |
| 7 | Freedom (Orlando) |
A | Davis | Davis | Torres | Brown | DNS | Tiger Invitational |