The England striker suffered minor ligament damage to his right ankle against Bayern Munich on Tuesday and is expected to be out for up to a month.
It is the nightmare scenario that was always likely as he played week in, week out and thundered into every challenge from first minute to last - and one that England boss Fabio Capello will monitor in forensic detail with his World Cup hopes so dependent on the 24-year-old's burly shoulders.
United will point to last weekend's 4-0 win over Bolton Wanderers as evidence that they can do well without him and Dimitar Berbatov, who scored two in that game, certainly seemed to enjoy the extra responsibility of leading the line.
But Chelsea are another kettle of fish and Rooney, with 34 club goals this season, will be hugely missed in what could be the pivotal game of the whole campaign.
"Obviously you want your full squad fit at this time of the season as every game is massive in the run-in so we are all hoping that Wayne is fit very soon," said Ryan Giggs.
With Michael Owen out for the season, United's striking options suddenly look thin and Italian teenager Federico Macheda, with just five minutes of league action under his belt this season, could be brought into the fold.
Last April he hit a vital injury-time winner against Aston Villa on his debut to halt a run of two defeats, then scored the winner against Sunderland to help United secure a third successive title.
"Young Macheda's had a terrible season with injuries but he made a return against Bolton and we were absolutely delighted," manager Alex Ferguson said of the striker.
"He's one of the most exceptional young strikers I've seen in my time at United. He has every chance of taking part in big matches."
In contrast, Chelsea will reap the benefit of their Champions League exit as they arrive refreshed for the game and with manager Carlo Ancelotti's only striking problem being whether to start with Didier Drogba, Nicolas Anelka or both.
Chelsea have beaten Portsmouth 5-0 and Aston Villa 7-1 in their last two games and Drogba watched the second of them from the bench as Frank Lampard took the honours with a four-goal haul.
Florent Malouda, with two goals in each of those games, is another player in top form and gives Ancelotti another attacking option from midfield as Chelsea seek their first Old Trafford win for five years.
That goal rush has edged their goal difference ahead of United's - plus-53 to plus-51 - with United top of the standings on 72 points to Chelsea's 71 with six games to play.
Hovering right behind them on 68 having lost ground with last weekend's draw with Birmingham City are Arsenal and a home win over Wolverhampton Wanderers will put them right back into the mix.
They will probably have to achieve that without Cesc Fabregas, Andrei Arshavin and William Gallas, all injured in Wednesday's Champions League draw with Barcelona.
Tottenham Hotspur, in pole position in the race for fourth place, have an awkward-looking trip to Sun
