"Gunslinger Girl" is a manga series set in a near-future Italy, where a government agency known as the Social Welfare Agency operates covertly to combat terrorism. The agency employs young girls who have suffered severe trauma, pairing them with adult male handlers to form specialized counter-terrorism units. These girls are physically enhanced through cybernetic implants and psychological conditioning, transforming them into highly efficient assassins. Despite their lethal capabilities, the girls retain childlike innocence and emotional vulnerability, creating a stark contrast between their humanity and their programmed roles as weapons.

The narrative primarily revolves around the relationships between the girls and their handlers, exploring themes of loyalty, dependency, and the ethical implications of using children as tools of violence. Each girl is assigned a handler who trains and oversees her missions, with the dynamics between them ranging from paternal care to cold professionalism. The protagonist, Henrietta, is paired with Jose, a handler who initially views their partnership as purely professional but gradually develops a protective bond with her. Other key characters include Rico and her handler Jean, whose relationship is marked by mutual respect and emotional distance, and Triela, who exhibits a strong sense of individuality despite her conditioning.

The plot unfolds through episodic missions interspersed with flashbacks that delve into the girls' tragic pasts and the circumstances that led to their recruitment by the agency. These missions often involve neutralizing terrorist threats, but the focus remains on the psychological and emotional toll on the girls and their handlers. The series also introduces antagonists, including rival factions and rogue agents, who challenge the agency's operations and force the characters to confront the moral ambiguities of their work.

As the story progresses, the girls' conditioning begins to erode, revealing their suppressed memories and emotions. This internal conflict is juxtaposed with external threats, such as the emergence of a powerful terrorist organization and internal corruption within the agency. The series culminates in a climactic confrontation that tests the limits of the girls' loyalty and the handlers' resolve, ultimately raising questions about the cost of sacrificing humanity for the sake of security.