| Team | Squad | Seed | Note | |
|---|---|---|---|---|
| 1 | Westside | A | 47.20 | |
| 2 | Hilliard | A | 54.47 | FHSAA 1A Region 1 | 
| 3 | Trinity Christian (Jax) | A | 54.48 | 54.48 Don Hall Invitational / St. John River Conference | 
| 4 | Calvary Christian (OB) | A | 54.59 | Ancient City Invitational | 
| 5 | Duval Charter | A | 55.61 | 29th Annual Chandra Cheeseborough Invitational | 
| 6 | Episcopal | A | 55.89 | |
| 7 | Bradford | A | 59.80 | Raider Fr/So Invitational | 
| 8 | Impact Christian | A | 1:02.84 | Ancient City Invitational | 
| 9 | Interlachen | A | NT | |
| 10 | Westside | B | NT |