Goals from Billy Sharp - his 27th of the season - and Jermaine Beckford sealed the three points.
It was a valuable success, as the game between second-placed Bristol City and third-placed Nottingham Forest finished 1-1.
Grant Holt had fired Forest ahead at Ashton Gate, but Bradley Orr levelled to keep City ahead of the visitors in the table.
The upshot of that for Scunthorpe is that they now hold a four point advantage over City, and an eight point lead over Forest with just six league games left to play.
Gary McDonald sealed a 1-0 win for Oldham over Yeovil to strengthen their play-off credentials, the result saw them leapfrog the Glovers in the table.
Blackpool's 2-1 win over Crewe keeps them in the final play-off spot, though they pulled away from seventh-placed Swansea - the Welsh side crashed 2-0 at Millwall.
Cheltenham climbed out of the drop zone with a 2-1 win over Leyton Orient.
Brentford look doomed though - they crashed 2-1 at Gillingham despite taking an early lead.
Rotherham also appear resigned to making the drop to League Two - they were beaten 2-0 at home by Bournemouth.
Huddersfield and Port Vale drew 2-2 at the Galpharm Stadium. Brighton were 2-0 winners at Northampton.