The heartfelt letter, which originally appeared in the 2007 book "Tupac Remembered", is reportedly on display at the Tupac Amaru Shakur Center for the Arts in Stone Mountain, Georgia, according to E! Online.
"When I was feeling at my worst, I knew I could put that Tupac tape in, and suddenly things weren't so bad. Thank you for giving us his spirit and yours! God bless you," read the letter by the "Lose Yourself" hitmaker.
Eminem, whose real name is Marshall Bruce Mathers III, also praised Afeni for being so warm to the rapper.
The letter also included a drawing Eminem created of the "All Eyez on Me" hitmaker with his signature bandanna.
"To Afeni, Your friend 4-ever1 Marshall," he signed the drawing.
Tupac was shot dead on September 13, 1996.