- Mandy and Cameron Klinefeld from season 4's "Assume Nothing" put the "unholy" back in Unholy Matrimony. A married couple united by a shared sexual fetish for rape and murder, the pair operate by abducting other husbands and wives, sexually torturing the women while making the men watch, with the ultimate goal of forcing them to Mercy Kill the women they love. They promise the husbands they can go free if they do this, but they're lying.
- Laura Gabriel, from season 12, is a ruthless arms smuggler and Big Bad Friend to Catherine Willows. Playing a meek and submissive wife to her sleazy private military firm head husband Mark, she uses this false personality to deflect blame off her and onto him as she funnels his company's weapons to the Mafia and the cartels. When the investigation starts getting too close to the truth, she hires employees of the company to murder her own lawyer and all of his staff; following this, she kills the assassins and burns their bodies beyond recognition in order to fake her own death and that of her right-hand man. She intended for an enraged Catherine to summarily execute her husband, and placed a contract on Catherine herself to clean up the last remaining loose end, not caring for her old friend in the slightest.
- Stewart Otis, first appearing in season 1's "Broken", is a pedophilic Serial Killer and Serial Rapist who targeted little girls around the ages of five to eight years. While acting as an authority figure, he would kidnap the girls to take to his home for him to rape and strangle to death before burying the bodies in is backyard. He has done this to eight little girls and only lamenting on his latest victim, Ruthie, because she died too quickly before he got a chance to "play" with her. He later appears in the season's finale "Body Count", where he escaped prison with the help of two fellow inmates, Hank Kerner and Randall Kaye. He then returns the latter's favor by kidnapping his daughter, Emma, taking her to a cabin where he not only raped her, but force her to partake in a film taunting Horatio Caine, the person who arrested him. He then tries to kill her before trying to find her cousin, Robyn, hoping to do the same thing to her.
- Henry Darius, from a CSI: Miami-CSI: New York crossover, is a Serial Killer-turned-Spree Killer known for shooting his victims in the head. As a child born from a mistress of a wealthy family, he always felt jealous that his half-sisters got more attention than he did, and as a result, became a murderer, with his first three victims looking like his sisters. First appearing in the season 4 Miami episode "Felony Flight", he causes a plane crash that kills two pilots and one U.S. Marshal. He then steals the marshal's gun and uses it to kill four college students and steal their car. He then travels to Miami University where he would kill two more college students after they told him about his half-sister, Alexa, and her whereabouts. He then goes to her location and kidnap her before shooting a man for his car and heading to New York City. When he arrives there in the season 2 NY episode "Manhattan Manhunt", he murders Alexa along with six college students, before heading off to kill his second half-sister Sarah. A narcissistic sociopath, with a violently-low impulse control, an excuse that doesn't come close to justifying his actions, and 20+ body count to boot, Henry Darius was among the worst that both CSI teams from Miami and New York had ever had to face.
- Python, real name Dante Wilkerson, is one of the world's most wanted hackers. A figure born with reptilian skin, he would run a website where he sells faulty guns and drugs in the hope of creating anarchy. When cornered by the authorities, he would brutally torture and kill any agents sent against him. He would also hack various government agencies and release the information to criminals. Becoming obsessed with Agent Avery Ryan, he would kidnap, torture and even, just to mess with her, would give drugs to Avery's friend. About to be captured, he threatened to use his tech skills to overdose the friend if they dont solve his riddles, but later attempts to kill her anyway. After failing, he tracks Avery down and kills the agents guarding her in the sick hope that if she kills him, she would become just like him. Python mixes cold sociopathy and vindictiveness and stands out as one of the most wicked of the franchise has ever produced.
- Josh Freely, from season 1's "Fly Away", is a brutal pedophile who uses his position, as a social worker working with emotionally unstable parents, to find new victims. Freely piles emotional abuse onto the parents, making them feel as though they're worthless, so they surrender their kids to him, and if that doesn't work, he's more than happy to fudge his own records so his superiors order the children removed. His abuse eventually reaches the point that one such mother attempts to kill herself and her daughter rather than let Freely take her.
- Jim Larkin, from the season 1 finale, "Lovers' Lane", is a slovenly glutton. Too lazy to even abduct victims himself, he instead badgers his abused and weak-willed son to do so for him, usually unattractive teenage girls he pretends to befriend. His assaults—including one murder—are brutal, reaching the point at which he murders one of his victims simply for calling him "disgusting".
- Roger Mulvaney, from season 3's "A Perfect Day", ruthlessly beats his wife and two daughters and uses his position as a police officer to get away with the abuse. Once his wife decides to leave him for another man he resolves that if he can't have them, nobody could. He kidnaps the three of them and makes his wife watch as he throws one of the girls—only four years old—from a tall bridge.
- "John Smith", real name unknown, from season 5's "The Road", is possibly the most horrific Serial Killer seen on the show. Disturbed since childhood, he described the sight of watching a woman drown while doing nothing to save her as the most beautiful thing he'd ever seen, and dedicated his adult life to replicating that "beauty". Abducting women who were perfectly happy with their lives, Smith brought them to special cellars where he sealed them off, watching as they went insane from isolation before finally leaving them there to starve.
- Season 3's finale and season 4's premiere had Emil Dornvald, a mercenary working for an African dictator. The missing person of the week is in love with a rebel opposing said dictator and knew where he and his friends were holed up. Dornvald cold-bloodedly murders the pregnant teenager she's been mentoring and is very fond of, sends her a picture on the girl's phone, and offers to stop killing her loved ones if she just tells him where the men are.
- Season 6's premiere had a conman preying on families who had adopted children from Africa; he'd claim to be the kid's birth father and guilt the parents into surrendering the kids to him, whereupon he'd take them and sell them on the black market. He also manipulated a Sudanese war orphan into helping him by claiming that he was only stealing the children to get them back to their birth parents, the young man having lost his own mother to militants. When the orphan finds out he's been had, he brutally stabs the guy to death and the FBI quite understandably decide to look the other way.
- Level 26: Dark Origins: This digital novel, by CSI creator Anthony Zuiker, features Sqweegel, real name unknown, the most prolific Serial Killer in history within the CSI world. Sqweegel is known for incredible preparation and his seeming randomness. Once he locks in on a victim, Sqweegel proceeds to murder them, often after torturing and raping them, with gender being irrelevant to his tastes. Sqweegel initiates the book's plot by sending in a homemade snuff film to the authorities, and proceeds to kidnap three teenagers who tried to illegally tried to buy beer before raping them with himself, a mop handle and a baseball bat. He burns six priests alive and attempts to torture the widows of 9/11 firefighters to death. His true enemy is the investigator Steven Dark, whose foster family was murdered by Sqweegel once Dark came close to catching him years ago. To punish Dark for cheating on his wife Sibby, Sqweegel had raped Sibby, possibly conceiving a child which Sqweegel plans to kill, along with raping and killing Sibby. With a body count numbering in the high hundreds and the belief he is an instrument of divine retribution, Sqweegel remains the most terrifying enemy of Dark's career, and is the only known serial killer to receive the classification Level 26 to designate a monster that was previously beyond human comprehension.