As the earliest example, Ultron having survived his destruction and rebuilding to become a new threat was certainly not expected when the fic began, made even worse as this is an Ultron who has learnt from his mistakes and is actively working to avoid repeating himself even as he is committed to a galactic version of his old goal.
Probably not completely unexpected, but chapter 23 ends with the Guardians of the Galaxy discovering an ancient ship on the outskirts of the galaxy, the following chapter depicting their own decision to enter the galaxy in search of another of Quill's half-siblings.
Plus, this was the first inclusion of characters from Star Wars: Knights of the Old Republic in the greater story, as HK-47 becomes a new member of the guardians.
Hector Ayala/White Tiger, mostly due to being a B-list hero who hasn't even appeared on the MCU yet, so his origin story was pretty unexpected. Same with his friend Jonathan "Jack" Hart and the first powered rogue he faces, Raymond Bloch/Ox.
Celeste Morne only appeared in a few specific comic storylines, but here she becomes a significant figure on Earth as its first known Jedi resident, a member of the Masters of the Mystic Arts, and Bucky's love interest (to say nothing of the enduring question of how she came to be on Earth in the first place).
Chapter 52 sees the introduction of the Ghost Rider, who normally has nothing to do with the Order of the Mystic Arts and their corner of the MU.
Darth Kreia and XoXaan, mostly because both were long dead by the time of the Clone Wars, so showing up as ghosts during the Dathomir Arc was very unexpected.
Barel Ovair, mostly because the character originally debuted during the Great Galactic War, almost 3000 years before the Clone Wars, so his appearance was rather surprising.
While Black Widow (2021) introduced an MCU version of the Taskmaster in canon, Chapter 62 features another Taskmaster in the form of an alien mercenary in the Star Wars galaxy.
During the Mortis arc, Thor appears in a Big Damn Heroes moment to help Anakin, Obi-Wan, Vision, T'Challa, and Spider-Man confront the Son after he nearly killed the Father and Ahsoka.
As if Thor alone wasn't enough, the same chapter sees Peter find what appears to be the Venom symbiote in the Son's area of Mortis.
And on top of all that, the following chapter sees Anakin face a certain Sith Lord in black armor, resulting in the first of what will likely be a few clashes between Anakin Skywalker and Darth Vader...
The Prime Celestial, Arishem, appearing in Mortis to deal with the Father, the Son and the Daughter, was a huge surprise to a lot of readers.
After Word of God stated that almost none of Phase 4 properties would appear in this story, no one was expecting not only the titular character, but the entire story of Shang-Chi and the Legend of the Ten Rings. Especially considering that Shang-Chi and Katy Chen are debuting a few years earlier than their canon counterparts.
Another surprise to be sure is the addition of Deliah Blue, a character from the legacy storyline of Star Wars Legends, a Zeltron companion of Cade Skywalker, descendant of Luke and Anakin. She appears as the first conscript in story, joining the 501st as an engineer.
The hints that Jorus C'Baoth and the Yuuzhan Vong will be joining the story are pretty surprising considering C'Baoth was several years dead by this point in Legends and the Vong didn't openly show up until forty five years after the Clone Wars
Unexpected as he was expected given him being listed on the Mycast page of this story, in chapter 86 we are met with Luthen Rael, a prominent figure in the show Andor which takes place a few decades after the Clone Wars.
Cosmo the Cosmonaut is met by the Guardians in Chapter 88. Though we saw her in the first Guardians movie, there was no indication or inclination that she would be making an appearance in the story proper. She came into prominence during the Christmas Special but was officially introduced in Phase 2 of the MCU.
It was hinted that he would appear in the story at some point, but no one expected Thrawn appearing in the Trandoshan moon during the Lost Padawan arc - and as an ally of Ahsoka, who is a bitter rival of his in Star Wars Rebels, The Mandalorian and her self-titled series.
Having an ancient Sith Lord, Darth Marr from the Old Republic MMORPG appear in a Holocron on Tython and offer assistance to Anakin Skywalker was certainly not something many would have expected to see in this story.
Even with the hints that the Great Web and the Spider-Verse would be involved, it's still a surprise to witness a Peter Parker who has been trained as a Jedi from a world where some of the Avengers are natives of the Galaxy Far, Far Away.
After IW!Peter and Jedi!Peter become involved in the events of Spider-Man: Into the Spider-Verse, while it was expected that the rest of the Spider-Gang would appear, their first encounter with one of the gang isn't Miles or any of the more 'direct' variants of themselves, but instead Peni Parker, who was basically just part of the group in the original film.
As the Spiders return to their worlds, a few further Spiders are featured; the Peter Parker and Miguel O'Hara of Spider-Man: Shattered Dimensions and the Spider-Man and Madam Web of Spider-Man: The Animated Series, with the implication that Madam Web will assist the newly-created "Alpha" in learning more about his powers and the others in forming a new variation of the "Spider Society" in Spider-Man: Across the Spider-Verse.
It's a small blink and you'll miss it moment but Katy Chen rescued Sintas Vel, a Kiffar Bounty Hunter who would eventually grow up to marry Boba Fett.
In the Season 4 trailer, the chapter ended on one of the most unexpected of reveals being Odin having met Revan in the past
Chapter 114 has the Sith Lords Darth Wyyrlok, Nihl and Maladi appear, ones who come from the future, not the past.
