| Lane | Team | Squad | Athlete 1 | Athlete 2 | Athlete 3 | Athlete 4 | Seed | Note |
|---|---|---|---|---|---|---|---|---|
| 1 | Roxana | A | Newton | Mitchell | McVey-Rismiller | Blas | 4:12.54 | |
| 2 | Highland | A | Wallace | Walker | Huelskamp | Brock | 4:00.45 | |
| 3 | Bethalto (Civic Memorial) | A | Silva | Fonrodona | Wooten | Eldridge | 3:55.00 | |
| 4 | Wood River (East Alton-W.R.) | A | Burch | Skinner | Burgess Jr | Loeffelman | 3:59.00 | |
| 5 | Jerseyville (Jersey) | A | Schultz | Talley | Judkins | Little | 4:10.00 | |
| 6 | Alton (Marquette) | A | Rochester | Fears | Nash | Kline | NT |