| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 2 | Lake Placid |
A | Holmes | Koontz | Bullard | Ritacco Jr. | DNS | Sebring Relay Meet 2025 |
| 3 | DeSoto County |
A | Tice | Wilkinson | Brown | Powers | 44.84 | Sebring Relay Meet 2025 |
| 4 | Avon Park |
A | Sheilds | Nelson | Kirkland | Naar | 44.50 | Sebring Relay Meet 2025 |
| 5 | Hardee |
A | Cherry | Dick | Myrick | Garner | 44.82 | Sebring Meet #2 2025 |
| 6 | Vanguard (LW) |
A | Khan | Lightborne | Seymour | Lacossade | 46.03 | Sebring Meet #2 2025 |
| 7 | Sebring |
A | DNF |