8 seed, finally earned the must-have, go-ahead service break with the help of a point in which the right-hander tumbled to his backside, scrambled back to his feet and hit a shot lefty.
He fashioned three break points from the next rally and took the second, the first break in 49 games.
Roanic, the 13th seed, required strapping to his right thigh in the first set and had treatment later in the match, but Isner deserved this victory.
Isner appeared to be struggling at times from 12-all to 18-all, but he held firm on his serve as Anderson, playing his second-consecutive five-set match, found a second (or third or fourth) wind.
Melichar and Peschke will play the victor of the second semifinal between No. 3 seeds Barbora Krejcikova and Katerina Siniakova of the Czech Republic and the unseeded pair of Alicja Rosolska of Poland and Abigail Spears of the U.S.
The semi-final was deep into the fifth set when the new record was set, although it is still a long way behind the 11 hours and 5 minutes it took Isner to beat Nicolas Mahut in the 2010 first round.
Isner and Raonic entered Wednesday night's quarter-final as the two leading players for aces in the tournament at 135 and 117 respectively. His 53 on Friday gave him 214 for the fortnight, beating the mark of 213 set by champion Goran Ivanisevic in 2001. Even Anderson himself used the match to call for change in format.
For the first time in the Open era, the semi-finals were made up exclusively of players over 30.
Nadal has 17 major titles, two at the All England Club.
Nadal, the 2008 and 2010 champion, trails Djokovic, the 2011, 2014 and 2015 victor, 26-25 in a rivalry which began at Roland Garros 12 years ago.