Brooke Mueller and her children with Charlie Sheen, 6-year-old twins, Max and Bob, are HIV negative.
Following the former Two and a Half Men star's Today show interview, Mueller's rep, Steve Honig issued a statement to ET.
"Brooke has been inundated with calls from friends and family," he said. "To put their minds at ease, Brooke can confirm that she and the boys are not HIV positive."
On Tuesday, Sheen told Matt Lauer that he told Mueller about his status, confirming that he immediately called her about the news and told her she should be checked.
Previously, a source tells ET that Denise Richards -- another ex-wife of Sheen -- and their two daughters, 11-year-old Sam and 10-year-old Lola, are HIV negative. The source added that Sheen got infected after he and Richards split in 2006.
Sheen also confirmed that Richards new of his status before making the announcement on Today.
Before Sheen's announcement, sources told ET on Monday that claims of the 50-year-old being HIV positive were true. According to reports, Sheen had known for some time and already confided with several individuals about it.
"What I can tell you is that if Charlie is indeed HIV positive and indeed owns it and comes forward, it can be a good moment,” Hollywood publicist Howard Bragman told ET on Monday. “I’m sure it’s a very hard personal moment. It takes a lot of courage on his part to talk about it.”
Meanwhile, former adult film star and Sheen's ex Bree Olson, spoke out publicly amid the HIV rumors.
"I, myself - have been to the gynecologist once a year since we split up and have always been tested across the board for everything and have come back clean across the board every time," Olson tweeted. "For all the 'news' sources out there, leave me alone. Call me Jon Snow. I know nothing."
Another source tells ET that Bree Olson is HIV negative.
ET has reached out to Sheen for comment. The actor, who is also father to a 30-year-old daughter, has not responded.
Watch more of Sheen's HIV admission below.