| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 2 | Mason Classical Academy | A | Bishop | Lindsey | Switken | Dix | 57.06 | Naples Quad Meet |
| 3 | St. John Neumann | A | Maguire | Seeber | Jean-Felix | Martin-Gonzales | 53.50 | |
| 4 | Estero | A | Guedouar | Joseph | Buckner | Hill | 48.79 | 48.79 5th Annual Jim Smith Invitational |
| 5 | Barron Collier | A | Aristizibal | Bongirno | Burger | Seaborn | 52.57 | FHSAA 3A District 12 |
| 6 | Bonita Springs | A | Dobson | Wolfson | Lipari | Baran | 55.11 | FHSAA 1A District 8 |
| 7 | Mariner | A | Simon-Lindsy | Marinell | Martinez | Rejes | NT |