Spies is the first satellite rider to start on pole since Monster Yamaha Tech 3 team-mate Colin Edwards at Shanghai, China, in 2008. The #11 is also the first rookie to start from pole since Jorge Lorenzo in 2008.
2010 title leader Lorenzo came closest to beating Spies, but the factory M1 rider - almost certain to ride alongside the Texan next season - was left 0.220sec behind at the chequered flag.
Lorenzo had knocked another American, Nicky Hayden, from pole with three minutes to go as he seeks his eighth win of the year on Sunday.
Hayden will start third after claiming his first ever front row for Ducati at the track where he finished third, marking his only Desmosedici podium to date, last year.
Team-mate Casey Stoner had been fastest in both practice sessions, but could only manage sixth - directly behind Repsol Honda riders Andrea Dovizioso and Dani Pedrosa - after being held up on his final fast lap.
Stoner appeared to get revenge on Hector Barbera for the obstruction by backing off soon after, delaying the Aspar rider and Pramac's Mika Kallio, on what would have been their last fast lap.
World champion Valentino Rossi crashed out with six minutes of the hour remaining.
The spill marked his second fall of the day and the Fiat Yamaha star will start just seventh on the grid. Rossi has struggled, by his standards, all weekend and was never in serious pole contention.
Fellow Italian Marco Simoncelli will start alongside Rossi in eighth - the San Carlo Honda Gresini rider also fell.
Completing the top ten will be Edwards and fellow veteran Loris Capirossi, of Rizla Suzuki.
Hiroshi Aoyama, making his MotoGP return from a fractured vertebra at Silverstone, qualified an impressive 13th out of the 17 riders on his Interwetten Honda.
Indianapolis Grand Prix - Qualification Results
1 Ben Spies (Yamaha Tech 3) 1:40.105
2 Jorge Lorenzo (Yamaha Factory) +00.220
3 Nicky Hayden (Ducati MotoGP) +00.231
4 Andrea Dovizioso (Honda HRC) +00.454
5 Dani Pedrosa (Honda HRC) +00.532
6 Casey Stoner (Ducati MotoGP) +00.559
7 Valentino Rossi (Yamaha Factory) +00.900
8 Marco Simoncelli (Honda Gresini) +00.987
9 Colin Edwards (Yamaha Tech 3) +01.127
10 Loris Capirossi (Suzuki MotoGP) +01.407
11 Alvaro Bautista (Suzuki MotoGP) +01.429
12 Marco Melandri (Honda Gresini) +01.518
13 Hiroshi Aoyama (Honda Interwetten) +01.526
14 Aleix Espargaro (Ducati Pramac) +01.544
15 Mika Kallio (Ducati Pramac) +01.751
16 Hector Barbera (Ducati Aspar) +01.791
17 Randy de Puniet (Honda LCR) +01.818