The Italian outfit recorded a 4-1 victory in the first leg, leaving Valencia with serious work to do if they are to progress to the final eight of the competition.
Tuesday's clash at the Mestalla will be played behind closed doors in an attempt to contain the spread of coronavirus, with Spain's health ministry revealing that the country is doing its best to avoid "large gatherings".
The atmosphere would bound to have been red-hot had supporters been allowed inside the stadium, but Valencia will not be able to draw energy from their fans as they attempt to turn the tie around.
Only on four previous occasions have a team managed to recover from a first-leg defeat by three or more goals to progress, and it is a huge ask for Los Che considering that they have only been victorious in one of their last seven matches in all competitions during a tough run of form.
Albert Celades's side were held to a 1-1 draw by Alaves in La Liga on Friday, leaving them down in seventh spot in the table. This is actually the first time since 2012-13 that Valencia have reached the last-16 stage of the Champions League, but they were finalists in both 1999-2000 and 2000-01.
The Spanish outfit have lost four of their six previous two-legged ties with Italian opposition; that said, they have been victorious in 13 of their last 19 European fixtures at the Mestalla.
The coronavirus outbreak means that Atalanta have not been in action since thumping Lecce 7-2 in Serie A at the start of the month. The Italian side will bring a four-match winning run into this contest, including the 4-1 success when the two teams locked horns in Italy on February 19.
Atalanta are again going well in the league this season with 48 points leaving them in fourth spot, but Gian Piero Gasperini's side were quite fortunate to progress from their Champions League group considering that they only picked up seven points from their six Group C matches.
The Italians were the only team to make their group-stage debut in 2019-20, but they have played 42 European fixtures throughout their history, reaching the European Cup Winners' Cup semi-finals in 1987-88.
Atalanta have lost five of their last six European matches, although it would be fair to say that La Dea have serious room for manoeuvre due to their excellent performance in the first leg.
Valencia Champions League form: LDWDWL
Valencia form (all competitions): LDLLWD
Atalanta Champions League form: LLDWWW
Atalanta form (all competitions): DWWWWW
As for Atalanta, Duvan Zapata was named on the bench in the first leg, but the 28-year-old, who has scored 12 times in 18 appearances this season, should be in the XI for this match.
Rafael Toloi is the Italian side's main fitness test due to a muscular problem, although the Brazilian is being tipped to return in time for Tuesday's contest.
The Italian side's squad is otherwise in good shape, though, meaning that there are not expected to be many alterations from the XI that started the first leg.
Valencia possible starting lineup:
Domenech; Wass, Diakhaby, Gabriel, Gaya; Torres, Parejo, Kondogbia, Soler; Rodrigo, Gameiro
Atalanta possible starting lineup:
Gollini; Palomino, Caldara, Toloi; Gosens, Freuler, De Roon, Hateboer; Pasalic; Zapata, Ilicic
We say: Valencia 1-1 Atalanta
It is difficult to know what type of match to expect due to the fact that there will not be any supporters inside the stadium. Valencia are capable of picking up a positive result on the night, but the Spanish outfit have left themselves with far too much to do in terms of turning the tie around.