Atalanta rely on Jeremie Boga and Ruslan Malinovskyi, while Fiorentina pick Krzysztof Piatek and Nico Goznalez for the Coppa Italia quarter-final.
It kicks off at the Gewiss Stadium in Bergamo at 17.00 GMT.
Coppa Italia Liveblog: Atalanta-Fiorentina and Juventus-Sassuolo
The winner will be decided after 90 minutes, extra time or penalties.
Gian Piero Gasperini is having his most difficult season on the Atalanta bench, crashing out of the Champions League and falling out of the top four in Serie A.
Duvan Zapata made his comeback last week, only to immediately suffer a relapse ruling him out for the rest of the season during a 2-1 home defeat to Cagliari.
Robin Gosens was sold to Inter and Roberto Piccoli loaned to Genoa, but Luis Muriel needs rest.
Therefore, new signing Boga is given his first start as a centre-forward, supported by Malinovskyi and Mario Pasalic.
La Dea were in the Final of the Coppa Italia last season and want to repeat the trick.
Fiorentina are in a similar position, selling top scorer Vlahovic to Juventus in the January transfer window.
Their first game since his departure was a 3-0 home defeat to Lazio, which did little to boost confidence.
Piatek is given the nod upfront, as he has an excellent Coppa Italia record after 10 goals in just eight games.
Nico Gonzalez and Riccardo Saponara complete the trident, with Lucas Martinez Quarta returning for his first game in defence since January 10.
Atalanta: Musso; Djimsiti, Demiral, Palomino; Hateboer, De Roon, Freuler, Zappacosta; Malinovskyi, Pasalic; Boga
Fiorentina: Terracciano; Odriozola, Martinez-Quarta, Igor, Biraghi; Castrovilli, Torreira, Maleh; Gonzalez, Piatek, Saponara