Mick Harford's side were unable to preserve their League status
Luton's relegation from the Football League was confirmed after they were held to a 0-0 draw by Chesterfield.
The result, coupled with Grimsby's win at Notts County, means that the Hatters cannot get out of the bottom two.
Luton made a brave attempt to remain in League Two, but a 30-point deduction for financial infringements proved too much to overcome.
Luton have been in the Football League for 89 years and won the League Cup in 1988, beating Arsenal in the final.
They won at Wembley only eight days ago, when Claude Gnakpa scored an extra-time winner to secure victory in the Johnstone's Paint Trophy in a fine match.
Glory at Wembley, with a record Hatters 40,000 fans in attendance, was a welcome reward for Luton after a season in which they were all but doomed to the drop before a ball was kicked.
Luton boss Mick Harford has guided his side to 13 victories so far this season, with 15 draws and 14 defeats - a record which would have left the club comfortably in mid-table, if the points deduction had not been in place.
But with four games left to play, the Hatters will have to start preparing for life outside the Football League.
And Harford, regarded as one of the club's best ever players, said the team could hold their heads high despite their relegation.
"I said to the players it's 100% not their fault," said Harford.
"The past incumbents of this club are to blame. The penalty points were a massive burden we could not overcome."
In Monday's match, Luton's Keith Keane volleyed over and Gnakpa missed a good chance in the first half.
Chesterfield's Allan Goodall, Gregor Robertson and Derek Niven all had good chances for the visitors.
Luton pressed after the interval as news filtered through of Grimsby's lead at Notts Country, with Sam Parkin looping a header on to the top of the crossbar.
Tommy Lee frustrated the hosts with fine saves from Parkin, Gnakpa and Asa Hall and Luton's day was summed up when Parkin sent a free header over in the 75th minute.
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