The Argentina international was sent off for dissent during a 2-0 league defeat to his parent club Juventus at the San Siro before the international break.
Higuain was furious when he was booked for a challenge on Medhi Benatia with seven minutes remaining and his protests earned him a second yellow card, before he left the pitch in tears.
Milan appealed the decision, hoping to see the ban reduced to one match, but the Federal Court of Appeal in Rome ruled on Friday that the original punishment would stand.
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The 30-year-old is Milan’s top scorer this season with seven goals in all competitions, having joined the club on loan from Juventus during the close season.
Milan are fifth in Serie A on 21 points, a point behind Lazio in fourth ahead of the two teams’ meeting at the Stadio Olimpico on Sunday.
(Reporting by Alasdair Mackenzie. Editing by Patrick Johnston and Toby Davis)
