The Pilgrims had most of the possession and the chances during the contest but they could not turn that into any goals and the visitors from London were ruthless in taking their opportunities.
The deadlock was broken in the 59th minute when Zian Flemming raced onto a pass from Duncan Watmore and slotted low past Argyle goalkeeper Conor Hazard.
Argyle pushed relentlessly for an equaliser but the finishing touch was not there for them, with goalkeeper Bartosz Bialkowski standing strong and making some good chances, while his defenders played their part in the rearguard action.
Then, deep into added time, Millwall grabbed a second goal through substitute George Saville to give a final scoreline which did not come close to accurately reflecting how the game had been played out.
There were two changes to the Argyle team from the one which started the 1-1 draw away to Hull City on Saturday as Kaine KeslerHayden and Mickel Miller were called up by Steven Schumacher.
Macaulay Gillesphey, who had made his first start of the season against Hull after a summer groin operation, and Joe Edwards were named among the substitutes instead.
With Edwards on the bench, the Pilgrims were captained by Jordan Houghton, who anchored a midfield trio alongside Finn Azaz and Adam Randell in a 4-3-3 formation.
Miller was at left-back for Argyle while Kesler-Hayden was on the opposite side of the defence, with Julio Pleguezuelo and Dan Scarr in the central positions.
Millwall included Plymouth-born former Ivybridge Community College student Ryan Leonard in their side against the club where he began his career. Former Argyle loan signing Joe Bryan was also in the Lions' line-up.
Neil Warnock, until recently the manager of Championship club Huddersfield Town, was among the packed Home Park crowd for the Tuesday night under the lights fixture.
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