Despite taking a second-half lead, Chelsea lost 2-1 at Leverkusen as coach André Villas-Boas suffered his fourth defeat in seven games and his side now have to beat Valencia in a fortnight to stay in the Champions League.
A header from Leverkusen defender Manuel Friedrich in the 91st minute saw the hosts complete a fairytale comeback as victory sent them both top of Group E and into the Round of 16.
Valencia's 7-0 rout of Genk leaves them second with Chelsea now third in the table and just one point separates the three teams.
After Ivory Coast star Didier Drogba's second-half strike had put Chelsea ahead, Swiss striker Eren Derdiyok came off the bench to equalise 93 seconds later before Friedrich's dramatic winner.
Chelsea must beat Valencia at Stamford Bridge to go through, while even if Leverkusen lose at Genk in a fortnight, they will at least finish second in the group.
The victory was extra sweet for Leverkusen's former Germany captain Michael Ballack, who orchestrated the hosts' attack against his former club and had a memorable 100th Champions League game.
"We tried to take a few more risks and felt we deserved the draw, but to go and take the victory, we are delighted," said the 35-year-old. Leverkusen coach Robin Dutt, in his first season with Bayer after joining from Freiburg, paid tribute to his players belief.
"The lads absolutely deserved to win and delivered a passionate performance," said Dutt. "We already gave ourselves a good chance by beating Valencia at home (in October), that speaks for the character and quality of this team. Against Genk, we will try and make sure we finish in first place."
German champions Borussia Dortmund, failed to keep the momentum rolling from their battling Bundesliga win over Bayern Munich at the weekend, and were skittled by two Robin van Persie goals in Arsenal's Emirates stadium.
The loss leaves them at the bottom of their table relying on slim mathematical chances of qualifying for the next round of the Champions League.