There were five changes to the side which drew 2-2 with Colchester United at the weekend.
Luke Daniels replaced Joe Anyon between the sticks, with Scott Wiseman coming in for Jordan Clarke at right wing-back. It was as you were for the three central defenders, as Jack King, Murray Wallace and Charlie Goode retained their place in the starting eleven, as did Scott Laird.
Sean McAllister continued to partner Neal Bishop in midfield, but there was a new look to the front three, with Hakeeb Adelakun, Tom Hopper and Kyle Wootton returning.
Former Iron man Sam Winnall was the first to have an effort on goal, inside of two minutes. However, despite testing the returning Daniels with a downward header, he was flagged for offside.
Mark Robins' side had the better of the opening exchanges and almost took the lead in the 10th minute. Adelakun, who was playing the supporting role to Hopper and Wootton, cut in off the right flank and drove a low shot, which Tykes 'keeper Nick Townsend scrambled to save.
The visitors included Arsenal loanee Dan Crowley in their starting eleven, and he went close in the 25th minute, but he angled his shot just wide of Daniels' post.
The men from Oakwell then had a golden opportunity to take the lead, as McAllister misplaced his backpass, leaving Marley Watkins with an empty net to tap in to. It took a miraculous recovery from Daniels to deny the forward, as he smothered the ball at his feet.
The Iron were keen to get back on the front foot quickly, and went close when Laird drove the ball low across the penalty area to the back post, where fellow wing-back Wiseman was waiting to tap home, but the chance was snuffed out by some good defending.
In a quiet opening period, chances were few and far between. Just before the break, Hopper worked his way into space on the edge of the area but dragged his effort wide, subsequently failing to test Barnsley stopper Townsend.
In fact, the five minutes before the break were probably the most positive for the United, with Adelakun drawing a save from Townsend after some tricky wing play.
That was to be the last action of the first half, as shortly after referee Ben Toner brought a halt to proceedings.
It was the Iron who made the better start to the second half and ten minutes after the restart they almost edged in front.
Hopper showed a terrific willingness to win the ball on the wing and just as it looked like he was ready to pull the trigger, he opted to square the ball to his strike partner Wootton, but underhit his pass, meaning the chance was gone.
That seemed to spark the second half into life, as the visitors went up the other end of the field minutes later and found the back of the net. Lewin Nyatanga rose highest to dispatch his header into the roof of Daniels' net after a well-worked short corner.
The Tykes could have doubled their lead in the 65th minute but, after climbing well, Winnall flicked a header just over the crossbar.
Manager Robins introduced Paddy Madden on the 68 minute mark as United looked to get back into the game. The Irishman took the place of Hopper up front.
The change seemed to do nothing to deter the visitors' search for a second and shortly after the lead would have been doubled if not for some more heroics from Daniels, who dived to his left to palm Winnall's curling effort away to safety.
Daniels could do nothing to prevent what would soon follow. As, in the 75th minute, Barnsley frontman Watkins fought doggedly on the touchline to win the ball and, when he did, he hit a low left footed shot which beat the Iron goalkeeper from close range.
Despite falling two goals behind, United continued to press for a goal and substitute Kevin van Veen went close in the 82nd minute, but his near-post header was just a fraction too wide.
Van Veen could have brought the Iron back into it late on, but he saw his low drive brilliantly tipped round the post for a corner by Townsend.
From the resulting corner however, Robins' side did manage to find a consolation goal, as Goode tapped home from close range after a scramble in the goalmouth.
It was too little too late for United as, shortly after, referee Toner called time on the game. The Iron rallied late on, but ultimately fell just short.
Scunthorpe United: Daniels, Wiseman, Goode, King (McSheffrey, 65), Wallace, Laird, McAllister, Bishop, Adelakun, Hopper (Madden, 68), Wootton (van Veen, 72).
Substitutes not used: Anyon, Syers.
Barnsley: Townsend, Smith, Nyatanga, Scowen, Winnall, Watkins, Rothwell (Digby, 81), Crowley (Wilkinson, 86), Mawson, Harris, Wabara (Bree, 55)
Substitutes not used: Roberts, Hourihane.
Attendance: 1,796 (398 away).
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