Zambrano left Wednesday night's game at Tampa Bay because of discomfort in his right shoulder. He was examined Thursday in Chicago by team orthopedist Dr. Stephen Gryzlo.
Zambrano, 8-3 with a 3.13 ERA in 16 starts for the NL Central leaders, had been set to pitch Tuesday night at home against Baltimore.
"I don't think there's a possibility at all," Cubs manager Lou Piniella said before the series finale against the Rays.
Zambrano was originally set to have an MRI exam Thursday. The Cubs decided to instead schedule an MRI arthrogram, a test in which dye is injected into the problem area to provide a more detailed look.
"I hope it's not serious," Piniella said. "I hope we find out what's wrong and we can take care of it in short order."
Left-hander Sean Marshall could be recalled from Triple-A Iowa to pitch against the Orioles in place of Zambrano.
Zambrano won a career-best 18 games in 2007.
In other injury news, Piniella said outfielder Jim Edmonds, who left Wednesday's game with a sore left arch, was available to pinch hit.
"I'm doing a lot better," Johnson said. "They're going to give me one more day. I think I'll be ready tomorrow. We'll see."
Alfonso Soriano remains scheduled to have his broken left hand re-examined Friday. The left fielder, who would like to return before the All-Star break next month, hopes to be cleared to increase his rehab program.