| Lane | Team | Squad | Seed | Note |
|---|---|---|---|---|
| 1 | Lakeland | A | 54.00 | |
| 2 | George Steinbrenner | A | 52.50 | |
| 3 | Haines City | A | 51.86 | 51.86 Polk County Championship 2019 |
| 4 | Dr. Phillips | A | 47.68 | 2019 FHSAA State Track & Field Championships |
| 5 | George Jenkins | A | 51.00 | |
| 6 | Celebration | A | 52.50 | |
| 7 | Plant City | A | 53.00 | |
| 8 | Mount Dora | A | 56.41 | St. Cloud Track Invitational |