The fifth-seeded Japanese player had to pull out before his second-round meeting with Santiago Giraldo of Colombia on Wednesday. Nishikori hurt his left calf at the grass-court tournament in Halle, Germany, two weeks ago and aggravated it during a five-set victory over Simone Bolelli on Monday.
Nishikori said he felt too much pain during warm-ups to even try to play his second-round match, which had been scheduled for Centre Court. He described the injury as a "small tear" but was set to undergo more tests to determine the severity.
"It was hurting just walking and running, so I decide not to play today," he said.
It's a disappointing exit for Asia's top men's player, who reached the U.S. Open final last year and had been set for a possible quarterfinal meeting with defending champion Novak Djokovic.
"I thought I was playing well on grass," Nishikori said. "So, very unfortunate to lose this chance."
Nishikori had to retire from the semifinals at Halle but came to Wimbledon hoping he'd be fit enough to play. He said the calf felt better before the match against Bolelli, but the injury flared up again on court.
"I thought it was really going to be OK. But last match in (the) fifth set I was hurting too much," he said. "I don't know how I fight through, but somehow I won. ... I thought it would get better in two days but, yeah, it didn't get better."
Nishikori now has a month off before his next tournament in Washington, D.C., and said he expects to be fully fit in time for the series of hard-court tournaments in the U.S.
"I think the injury should be okay for next tournaments," Nishikori said.