Diego Simeone's side visit the Santiago Bernabeu on Sunday in La Liga, aiming to complete a hat-trick of victories over their bitter rivals and wrestle their way into the title race.
Atletico inflicted Madrid's only top-flight defeat of the season in September, with a 3-1 home win that shook Carlo Ancelotti's team.
Madrid responded with a run of 21 matches without losing, brought to a juddering halt by Atletico again in the Copa del Rey last 16 in January.
The Rojiblancos romped to a 4-2 extra-time victory to gain revenge for Madrid's Spanish Super Cup semi-final win against them the week before in Saudi Arabia.
Another win for Atletico on Sunday would mean more than taking the derby honours - it would help revive their title hopes.
Real Madrid lead second place Girona by two points and Atletico, third, by 10.
Simeone insisted that the previous matches held little relevance ahead of the third clash with Madrid in under a month.
"It's a different game, as always in the derby," said the coach after Atletico secured a 2-1 win over Rayo Vallecano on Wednesday.
Atletico striker Memphis Depay has hit a rich vein of form ahead of the derby. The Dutch forward has scored in his last three matches.
"Memphis is working very well in the box, he's very powerful, very strong," said Simeone.
"We have to look after him, we need him and I hope he continues like this because the team needs these performances he's putting on now."
Both sides have problems in defence, with Atletico missing Jose Gimenez and Cesar Azpilicueta, while Madrid are without long-term absentees Eder Militao and David Alaba, as well as goalkeeper Thibaut Courtois.
Ancelotti said he did not think Atletico had his team's number after they put four past Madrid in the Copa del Rey.
"I don't think so, because a few days ago we beat them," pointed out the Italian coach.
"Atletico have very dangerous players, who can score goals against you... the last two games we've played against them, we could have done better.
"We've also scored a lot against them too, and against Atletico things aren't so simple."
The Italian was able to rest some players as Madrid eased to a 2-0 win over Getafe on Thursday night, knowing on Sunday they have a chance to knock Atletico out of the title race.
Player to watch: Ilkay Gundogan (Barcelona)
The Barcelona midfielder has not hit the heights of his finest days at Manchester City but is improving despite the team's struggles.
Gundogan's delivery contributes a large part of Barcelona's attacking edge and he will aim to keep the champions' withering title defence alive at Alaves.
Key stats
10 - Athletic Bilbao goalkeeper Unai Simon leads La Liga in clean sheets.
14 - Getafe's Borja Mayoral has pulled level with Real Madrid's Jude Bellingham and Girona's Artem Dovbyk as the league's top goalscorer.
18 - Villarreal midfielder Alex Baena has played more through balls than any other player.
