Scunthorpe ended Barnsley's FA Cup hopes with a fine 2-0 win at Glanford Park and now face a long trip to Premiership side Portsmouth.
Iron boss Brian Laws was delighted with his side's performance and said: "It's been a fantastic night and I am so delighted for the players.
"We were the underdogs, and they are an outstanding side in the second division.
"We asked the players to come out fighting and they took it on board and showed great character.
"I thought we wanted it more than they did. We've played better football and not won but we had the will to win." Gudjon Thordarson, Barnsley's under-pressure boss said: "We are very disappointed. We knew it would be a tough game, we didn't compete and were never good enough." Barnsley opened strongly and Tony Kay directed a third minute header wide and two minutes later Steve Hayward's free-kick caused no problems for the Iron's rearguard.
Scunthorpe shook the Tykes when they took a 14th minute lead as Lee Ridley hammered a free-kick in from the left and Steve Torpey rose above the visitors defence to flick a header home.
Torpey was a pivotal figure as the second division side began to force the pace, managing to get in several telling headers to clear attacks.
The nearest Barnsley came to equalising came when Jacob Burns got in a point-blank shot which Tom Evans did well to stop and then the keeper blocked an effort from Isaiah Rankin.
Scunthorpe almost grabbed a second on the restart, but Craig Ireland came in with a superb tackle to rob Steven MacLean in full flight.
Ireland then found himself at the other end and he turned Hayward's cross onto the woodwork.
That was almost the end for the Tykes as Scunthorpe hit them on the counter, makeshift striker Jamie McCombe curled home a superb right-foot shot from deep on the right in the 74th minute to take the wind out of Barnsley's sails.