Eddie Johnson, Perry Kitchen and Luis Silva scored for DC United, as the capital club won at home for the first time in three MLS fixtures, moving within a point of Eastern Conference leaders Sporting Kansas City.
Chivas hit back with six minutes remaining when Erick Torres scored from the spot but it meant little as the men in red and white stripes suffered defeat in MLS for the first time since May.
Johnson opened the scoring with a penalty in the 25th minute, following Oswaldo Minda's trip on Chris Rolfe in the box, while Kitchen doubled DC's lead 13 minutes after half-time with an opportunistic striker after Chivas failed to clear a cross into their defensive area.
Silva's third goal for the hosts was the best of the game, as he peeled off the shoulder of his marker to get on the end of Christopher Korb's through ball and then finished from a tight angle past visiting goalkeeper Dan Kennedy.
Steve Birnbaum's push on Mauro Rosales in the penalty area in the 82nd minute gave Chivas an opening with Torres converting from the spot.
But Wilmer Cabrera's side were unable to pull off an incredible comeback to stretch their MLS unbeaten run to six matches.
The defeat leaves Chivas in eighth in the Western Conference, one point behind seventh-placed Portland Timbers.
United's win took them to within striking distance of reigning MLS champions Sporting, with Ben Olsen's team having a game in hand on Kansas City.
