Four points over the Easter period and a third consecutive clean sheet was not enough as United's slim play-off chances evaporated at Tranmere Rovers.
A 0-0 draw, combined with Peterborough United's result on Easter Monday ensures the Blades cannot mathematically make the end of season play-offs.
For United's 54th competitive fixture of the campaign, boss Clough freshened up the starting XI - he made no less than five changes with Hill, Porter, McGinn, Davies and 18-year-old Dimaio all in from the start, whilst Collins took the captain's armband with Doyle rested.
McGinn appeared the more advanced of the midfielders in the early stages but Tranmere made the better start in terms of territory. In the opening 15 minutes the home side had two penalty appeals turned down and Akpa Akpro and Cassidy tried their luck from distance. The best effort of the opening exchanges, though, arrived on 14 minutes when Lowe held off Collins and saw his chip hit a post - the rebound also fell to the Tranmere captain but his second effort was straight at Howard.
United's early endeavours saw Davies whip in a couple of teasing crosses, Dimaio hit a half-volley from 20 yards and, on 21 minutes, Porter lost his marker but headed over a right-wing corner.
Another Dimaio half-volley, again the result of a Brayford throw, was closer but still not on target and the better chances were still falling to Tranmere. Around the half-hour mark, Lowe curled wide after a slip by Collins and Howard was called upon to push over a Brown shot.
As half-time approached, United switched to a 4-4-2 system, Murphy joining Porter in attack, whilst Tranmere were forced into their second change through injury, but neither team were able to break the deadlock before the interval.
It appeared that the first half display prompted boss Clough to make a double change at the break. Doyle and Baxter took over from McGinn and Porter and the Blades made a better start but were unable to test goalkeeper Fon Williams.
Maguire was the first in the referee's book, for halting the run of Cassidy, and on 55 minutes Taylor went close to converting Power's free-kick with a flicked header. Cassidy went close soon after but his shot from the edge of the box was blocked by a number of yellow shirts.
United's shouts for a penalty were ignored on 66 minutes despite Murphy staying down under Taylor's challenge and moments later Clough was forced into his third and final change when Freeman took over from Maguire with Hill moving to a central position.
Brayford fired wide on the run and a Power shot was hammered at Hill as the game began to open up in the final 20 minutes. Another flicked header from Taylor was wide, this time from Brown's delivery.
On 84 minutes Baxter cut in from the left, held off his marker, and forced a full-length save from Fon Williams in front of the Blades fans and Brayford headed wide the resulting corner. A minute later Cassidy failed with what looked the easiest chance of the day, missing a header across goal from a matter of yards and then Doyle blocked from Lowe as two minutes of injury-time was announced.
The game remained open late on but neither side were unable to strike that vital blow which would have won the game.
Tranmere: Fon Williams, Holmes, Taylor, Lowe, Cassidy (Stockton 90), Power, Jennings, Brown, Ridehalgh (Rowe 41), Akpa Akpro (Kirby 19), Pennington.
Unused: Morris, Bolland, Arthurworrey, Mooney.
Blades: Howard, Brayford, Maguire (Freeman 67), Hill, Porter (Baxter 45), McGinn (Doyle 45), Collins, Coady, Davies, Murphy, Dimaio.
Unused: Flynn, Paynter, Long, Kennedy.