Scunthorpe claimed a much-needed Coca-Cola Championship victory after Paul Hayes came up with a last-gasp winner at Glanford Park.
The hosts finally broke the deadlock when Matt Sparrow pulled back for Gary Hooper to score with a clever flick.
Barnsley pulled themselves level when poor defending from Rob Jones allowed Daniel Bogdanovic to fire home.
But with a draw looming, the ball fell inside the box to Hayes who curled beyond Luke Steele to claim the points.
The result was harsh on Barnsley who deserved a point after a evenly-balanced game.
Filipe Teixeira and Bogdanovic both wasted good chances with their heads for the Tykes, while Martyn Woolford and Hooper had further opportunities for the hosts.
Scunthorpe manager Nigel Adkins: "We're delighted we got the victory.
"It was a great finish by Paul Hayes, we built up some great pressure at the end there with a series of corners.
"Great credit to the players, the character that was on show after the defeat at the weekend was brilliant."
Barnsley boss Mark Robins:
"If you don't take your chances then unfortunately you're not going to get a point. We created a fair few, missed them and so we got punished.
"I expected to get at least a point from this game, if not the three.
"I'm really gutted we didn't get a point but on the balance of play I think we should have got all three."
90:00+5:59 The final whistle goes and the game is over.
90:00+5:35 Effort on goal by Gary Hooper from outside the penalty box goes harmlessly over the bar.
90:00+4:16 Assist on the goal came from Grant McCann.
90:00+4:16 GOAL - Paul Hayes:Scunthorpe 2 - 1 Barnsley Paul Hayes grabs a goal from just inside the area to the bottom right corner of the goal. Scunthorpe 2-1 Barnsley.
90:00+3:56 Corner taken left-footed by Grant McCann,
90:00+1:47 Inswinging corner taken right-footed by Michael O'Connor from the left by-line, Paul Hayes concedes a free kick for a foul on Carl Dickinson. Direct free kick taken by Luke Steele.
89:19 Unfair challenge on Martyn Woolford by Stephen Foster results in a free kick. Grant McCann produces a strike on goal direct from the free kick, Daniel Bogdanovic gets a block in. Inswinging corner taken by Michael O'Connor from the left by-line, Headed effort from inside the area by Rob Jones misses to the right of the target.
84:10 Daniel Bogdanovic delivers the ball, Rob Jones manages to make a clearance. Outswinging corner taken right-footed by Filipe Teixeira, Darren Moore gives away a free kick for an unfair challenge on David Mirfin. Joe Murphy restarts play with the free kick.
83:47 Andrew Wright takes a shot from 18 yards. Save by Luke Steele.
82:24 Centre by Paul Hayes.
81:32 The ball is crossed by Martyn Woolford.
80:09 Foul by Matthew Sparrow on Adam Hammill, free kick awarded. Carl Dickinson produces a cross, Filipe Teixeira takes a shot from just inside the penalty box which clears the bar.
79:56 Unfair challenge on Adam Hammill by Matthew Sparrow results in a free kick. Carl Dickinson takes the direct free kick.
77:54 The assist for the goal came from Bobby Hassell.
77:54 GOAL - Daniel Bogdanovic:Scunthorpe 1 - 1 Barnsley A goal is scored by Daniel Bogdanovic from deep inside the penalty area low into the middle of the goal. Scunthorpe 1-1 Barnsley.
76:26 The offside flag is raised against Gary Hooper. Indirect free kick taken by Carl Dickinson.
75:11 Gary Hooper challenges Stephen Foster unfairly and gives away a free kick. Free kick taken by Carl Dickinson.
74:16 Free kick taken by Luke Steele.
74:16 Booking The referee cautions Grant McCann for unsporting behaviour.
74:08 Grant McCann challenges Filipe Teixeira unfairly and gives away a free kick.
Correction - 70:26 Substitution Andrew Wright is brought on as a substitute for Michael Raynes.
Correction - 70:26 Substitution Sam Togwell leaves the field to be replaced by Grant McCann.
Correction - 70:26 Substitution Michael O'Connor comes on in place of Josh Wright.
Correction - 70:26 Outswinging corner taken by Filipe Teixeira.
70:26 Substitution Michael Raynes leaves the field to be replaced by Andrew Wright.
70:26 Substitution Grant McCann is brought on as a substitute for Sam Togwell.
70:26 Substitution (Scunthorpe) makes a substitution, with Michael O'Connor coming on for Josh Wright.
70:26 Outswinging corner taken from the right by-line by Filipe Teixeira.
69:49 The ball is sent over by Adam Hammill, save by Joe Murphy.
68:15 Gary Hooper takes a shot. Luke Steele makes a save.
68:15 Substitution Andy Gray leaves the field to be replaced by Jay Rodriguez.
68:15 Martyn Woolford produces a right-footed shot from just outside the penalty area that goes harmlessly over the target.
67:52 Centre by Matthew Sparrow, save made by Luke Steele.
67:14 Adam Hammill produces a cross, Daniel Bogdanovic takes a shot. Joe Murphy makes a comfortable save.
66:11 The ball is sent over by Matthew Sparrow, save by Luke Steele.
65:17 The assistant referee flags for offside against Paul Hayes. Luke Steele takes the free kick.
64:46 Free kick awarded for a foul by Hugo Colace on Sam Togwell. Free kick taken by Joe Murphy.
64:00 The referee blows for offside. Luke Steele takes the indirect free kick.
62:08 The ball is delivered by Adam Hammill, clearance made by Josh Wright.
61:17 Effort on goal by Daniel Bogdanovic from 18 yards. Save made by Joe Murphy.
60:04 Substitution (Barnsley) makes a substitution, with Daniel Bogdanovic coming on for Anderson De Silva.
60:04 The ball is delivered by Adam Hammill, clearance by Rob Jones. Centre by Bobby Hassell, Close range headed effort by Filipe Teixeira goes wide of the left-hand upright.
58:37 The offside flag is raised against Paul Hayes. Indirect free kick taken by Stephen Foster.
57:51 Shot from 20 yards by Paul Hayes. Carl Dickinson makes a save. Outswinging corner taken left-footed by Martyn Woolford, clearance by Andy Gray.
57:02 Gary Hooper takes a shot. Save made by Luke Steele. Long range shot by Sam Togwell goes wide right of the target.
56:26 Shot from just outside the box by Martyn Woolford misses to the right of the target.
53:39 A cross is delivered by Filipe Teixeira, save by Joe Murphy.
51:45 Adam Hammill is flagged offside by the assistant referee. Joe Murphy takes the free kick.
49:56 Assist on the goal came from Matthew Sparrow.
49:56 GOAL - Gary Hooper:Scunthorpe 1 - 0 Barnsley Gary Hooper grabs a goal from inside the area to the bottom left corner of the goal. Scunthorpe 1-0 Barnsley.
49:51 The ball is sent over by Matthew Sparrow,
48:02 The assistant referee signals for offside against Andy Gray. Rob Jones takes the indirect free kick.
47:30 The ball is swung over by Gary Hooper, save by Luke Steele.
46:43 A cross is delivered by Sam Togwell, Carl Dickinson makes a clearance. Josh Wright delivers the ball in from the corner, blocked by Carl Dickinson.
45:01 The referee gets the second half started.
Half Time
45:00+2:15 The half-time whistle blows.
45:00+1:45 The ball is swung over by Filipe Teixeira.
43:58 Rob Jones gives away a free kick for an unfair challenge on Filipe Teixeira. The free kick is swung in right-footed by Hugo Colace.
43:13 A cross is delivered by Adam Hammill, Shot from deep inside the area by Hugo Colace clears the bar.
42:20 Paul Hayes crosses the ball, save made by Luke Steele.
41:58 Gary Hooper sends in a cross, clearance made by Stephen Foster.
40:38 Adam Hammill produces a right-footed shot from outside the box and misses left.
40:02 Bobby Hassell delivers the ball, clearance by Rob Jones.
37:56 Nathan Doyle produces a volleyed left-footed shot from 35 yards and went wide right of the target.
37:26 Corner taken by Filipe Teixeira from the right by-line played to the near post, clearance made by Josh Wright.
36:16 Unfair challenge on Nathan Doyle by Matthew Sparrow results in a free kick. Direct free kick taken by Luke Steele.
34:42 Substitution Kieran Trippier leaves the field to be replaced by Bobby Hassell.
33:11 Filipe Teixeira produces a drilled right-footed shot from 20 yards. Blocked by Rob Jones.
32:35 David Mirfin concedes a free kick for a foul on Andy Gray. Carl Dickinson delivers the ball from the free kick left-footed from left channel, Paul Hayes makes a clearance.
31:52 Shot on goal by Adam Hammill from just inside the area goes over the target.
31:39 Filipe Teixeira crosses the ball, clearance made by Rob Jones.
31:00 The ball is sent over by Filipe Teixeira, Michael Raynes makes a clearance. Outswinging corner taken right-footed by Adam Hammill played to the near post, Josh Wright manages to make a clearance.
30:24 The ball is swung over by Adam Hammill, Michael Raynes manages to make a clearance.
29:28 Filipe Teixeira takes a shot. Marcus Williams gets a block in. Outswinging corner taken by Filipe Teixeira from the right by-line.
28:43 Corner taken right-footed by Filipe Teixeira from the left by-line, Paul Hayes manages to make a clearance.
28:12 The assistant referee flags for offside against Gary Hooper. Free kick taken by Stephen Foster.
27:46 Effort on goal by Martyn Woolford from just outside the area goes harmlessly over the target.
26:44 Gary Hooper fouled by Stephen Foster, the ref awards a free kick. The free kick is delivered right-footed by Josh Wright from left wing, Luke Steele makes a save.
26:00 Free kick awarded for a foul by Darren Moore on David Mirfin. Joe Murphy takes the direct free kick.
25:30 Unfair challenge on Hugo Colace by Sam Togwell results in a free kick. Carl Dickinson takes the free kick.
24:01 Nathan Doyle concedes a free kick for a foul on Matthew Sparrow. Josh Wright crosses the ball from the free kick right-footed from right channel, save by Luke Steele.
22:05 Shot from just outside the box by Gary Hooper misses to the right of the goal.
21:32 Anderson De Silva concedes a free kick for a foul on Martyn Woolford. Free kick taken by Josh Wright.
20:40 Handball by Darren Moore. Josh Wright takes the free kick. The ball is sent over by Martyn Woolford, Luke Steele makes a save.
19:05 Handball by Filipe Teixeira. Josh Wright takes the free kick.
17:41 Hugo Colace concedes a free kick for a foul on Marcus Williams. Free kick taken by Josh Wright. Darren Moore fouled by Rob Jones, the ref awards a free kick. Luke Steele takes the free kick.
16:47 Gary Hooper takes a shot. Save by Luke Steele.
16:30 Foul by Adam Hammill on Sam Togwell, free kick awarded. Joe Murphy takes the free kick.
15:49 Filipe Teixeira delivers the ball, clearance by Josh Wright.
14:32 The ball is crossed by Anderson De Silva, save by Joe Murphy.
13:45 Paul Hayes challenges Carl Dickinson unfairly and gives away a free kick. Carl Dickinson restarts play with the free kick.
12:57 Effort on goal by Martyn Woolford from 18 yards. Blocked by Kieran Trippier. Corner taken by Josh Wright from the right by-line, save made by Luke Steele.
11:07 Free kick awarded for an unfair challenge on Josh Wright by Nathan Doyle. Josh Wright restarts play with the free kick.
9:24 Unfair challenge on Adam Hammill by Sam Togwell results in a free kick. Carl Dickinson takes the direct free kick.
6:41 Kieran Trippier challenges Martyn Woolford unfairly and gives away a free kick. Josh Wright takes the free kick.
4:27 Anderson De Silva takes a shot. Michael Raynes gets a block in. Filipe Teixeira produces a right-footed shot from just outside the area that goes harmlessly over the target.
1:54 Anderson De Silva takes a shot from long range which goes wide right of the target.
1:20 Martyn Woolford delivers the ball, Kieran Trippier gets a block in.
0:00 The game gets underway.
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