Sometimes student, sometimes scientist and sometimes photographer, Peter Parker is a full-time super hero better known as the web-slinging and wall-crawling Spider-Man. As an orphaned child, Peter was raised by his Uncle Ben and Aunt May. At a science expo, Peter was bitten by an errant radioactive spider which granted him an array of arachnid-based powers. He initially became Spider-Man to use his powers as an entertainer, growing so conceited he did not bother stopping a passerby burglar. In a twist of fate, the same burglar wound up killing Peter's Uncle Ben, leading him to realize that he needed to use his powers responsibly. From then on, Spider-Man became a crime-fighting vigilante.

Miles Morales is a teenager from Brooklyn who was bitten by a Genetically Altered Spider and became Earth-1610's Spider-Man following the supposed death of that reality's Peter Parker.After the Multiverse was destroyed and subsequently rebuilt, Miles Morales, along with his friends and family, were transported to Earth-616 and rewritten into that world's history.

Jessica Drew has acquired bizarre powers as a consequence of her enigmatic life story that led her to become the dark angel vigilante and private investigator known as Spider-Woman. Through means provided by the High Evolutionary, an ill Drew was experimented on by her own father as an infant in a process that added the genome of a spider to her own. This granted her increased strength and agility, the power of generating venom blasts, and the ability to release disruptive pheromones. As an additional consequence, she lost many years of her life trapped in a coma. Revived as an adult, she was soon manipulated by Hydra into becoming one of their operatives. After attacking S.H.I.E.L.D. and Nick Fury, she realized she served corrupted forces and defected.

Flash Thompson is a bully-turned-selfless-soldier better known as Agent Anti-Venom. Initially the high school tormentor of Peter Parker, whose secret alter-ego of the super hero Spider-Man also happened to be Flash's biggest idol, Thompson grew out of his abusive behavior and befriended Peter in college. Flash joined the Army and was stationed in Southeast Asia,but had trouble finding a place to fit when he returned to the States.Flash spiralled into depression and combated alcoholism,an addiction that was exploited by the villainous Norman Osborn to involve him in a car crash. This left Flash in a near-vegetative state after waking up from a coma.Flash eventually regained his full faculties, and became a gym teacher at his old school.

In an average first day at school, she saw her friend Lynn Sakura being bullied by an older boy. Anya defended her friend, sparking a fight. A teacher broke their conflict up, but that night the two went down to a bridge in a nearby park to resume their battle. When she got there, Anya discovered a stranger being attacked by a strange group. This Sisterhood of the Wasp attempted to kill the man, but Anya leaped in the way of the killing blow. The site was allegedly a sacred spot. The man, Miguel Legar, was the sorcerer of the Spider Society, a secret group dating back for hundreds of years. Recognizing that Anya had the ability to become the new Hunter of the Spider Society, he transferred some of his power to her to save her life. As Legar's partner, Anya visited the WebCorps Headquarters, a shell company used to hide the Spider Society's actions, and became suddenly involved in the war against the Wasps. When Legar got caught in a bad situation when doing reckon work, WebCorps enlisted Anya to rescue him. During the confrontation, Anya was shot in her arm. The wound became a spider-shaped metallic tattoo. At WebCorps, she was informed of her birthright role as a chosen initiate for the Spider Society.

Ben Reilly is a genetic clone of Peter Parker, the super hero Spider-Man, who went on to establish his own identity as the Scarlet Spider.Ben was created by Miles Warren, the super villain known as the Jackal, to torment the hero, and seemingly died in their first encounter.The clone survived and exiled himself for five years, only returning to New York after learning that Parker's Aunt May was sick. After making amends with Peter, Ben fought alongside him as the Scarlet Spider.For a brief time, Ben and Peter were tricked by Norman Osborn via Seward Trainer to believe Ben was the real Peter Parker. Peter left New York with his wife Mary Jane Watson, leaving Ben to pick up the mantle of Spider-Man. During a subsequent confrontation against Osborn as the Green Goblin, Ben was impaled and died in Peter's arms, also discovering that Peter being the clone was a fabrication.

After being bitten by a genetically-engineered spider, Gwen was granted arachnid-like super-powers, and started a career as a crimefighter, dubbed by the media as "Spider-Woman." She was given a costume and a set of Web-Shooters by retired crimefighter Janet van Dyne. Gwen spent most of her early adventures focused on exploiting and maintaining her newfound attention more than helping those in need; however, Gwen's behavior changed after her father expressed he believed Spider-Woman could easily help people.