Botswana's Amos strode home in one minute, 43.63 seconds as Kenyan Rudisha, on the track for the first time in a year after a lengthy knee injury, fell back to seventh in 1:44.87.
"I just wanted to run my race," said Amos. "I think the world record can go this year."
Rudisha set that mark of 1:40.91 in winning the London Olympics and he admitted more training will be required if he is to get back to top form.
"It has been like a crash programme for me," said the soft-spoken Kenyan, who led or ran near the front for much of the race.
"It was tough for me but I was satisfied with my time. To run 1:44 for the first race after missing for such a long time I think it's good."
Ethiopian world champion Mohammed Aman claimed second in 1:43.99 with Sudan's Abubaker Kaki third in 1:44.09.
Ten other season-leading marks tumbled on a sun-splashed afternoon, none more impressive than Djiboutian Ayanleh Souleiman's 3:47.32 mile - the fastest ever over the distance on U.S. soil.
Another highlight was the 400 metres battle between Grenada's Olympic gold medallist James and American world champion Merritt, who came home side-by-side in the first sub 44-second runs of the year.
Both were timed in a photo-finish at 43.97 seconds with James handed the victory.
"He dipped and I didn't quite dip as well," said Merritt.
American world silver medallist Justin Gatlin continued his unbeaten 100 metres season with a wind-assisted run of 9.76 seconds, while French indoor world pole vault record holder Renaud Lavillenie claimed his specialty with a best of 5.80 metres.
World class fields in the women's 200m and men's 110m hurdles, however, were victims of upsets.
Little known American Tori Bowie, who learned only two days ago she would be in the meeting, shocked U.S. Olympic champion Allyson Felix and Jamaican world winner Shelly-Ann Fraser-Pryce in the 200 in the year's fastest time, 22.18 seconds.
Felix was third in 22.44 and athlete of the year Fraser-Pryce, who is nursing a lower body injury, was dead last in 23.06.
French indoor silver medallist Pascal Martinot-Lagarde surprised Jamaica's Hansle Parchment and world champion David Oliver of the United States for the high hurdles victory in 13.13 seconds.
World record holder Aries Merritt withdrew with an aggravated hamstring suffered in practice on Friday.
Kenya's Caleb Mwangangi Ndiku (men's 5,000 metres) and Hellen Obiri (women's 1,500m), Ethiopian Sofia Assefa (women's steeplechase), Jamaican Kaliese Spencer (women's 400m hurdles) and Russian Olympic champion Anna Chicherova (women's high jump) also had season-leading performances.
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