Luke has pale skin, long blonde hair tied with a white ribbon at the end, and green eyes. He wears silver pince-nez glasses and the same style of military jacket as his brother Jan, although Luke's is white with gold lining. In the manga, his jacket is black with white lining. He also wears white suit pants, white gloves, a white suit jacket over his jacket; in the OVA, a white overcoat draped over his shoulders is added. He later loses this overcoat during the Valentine brothers' invasion of Hellsing headquarters.
Despite being Jan Valentine's brother, the two are polar opposites. While Jan is socially loud, obnoxious, unpleasant and murders with glee and sadism Luke is far less talkative, usually calm and kills casually. Luke likes to fancy himself as the one who will bring about Alucard's demise, believing that because of his relatively advanced powers that he was destined to surpass Alucard in every way. He believes that he has more than Alucard's powers at his own disposal, and believes Alucard to be just no more powerful than an artificial vampire like himself. When he discovers this to be far from the case, he is shown to be cowardly, yet still tries to put down Alucard, foolishly claiming him to be nothing more than "a dog."
He and his brother Jan are both sent to attack the Hellsing headquarters.
Luke is later sent to battle Alucard, and he believes himself to be more than a match for Alucard. He soon finds that this is far from the truth, being soundly beaten by Alucard, who pleads with Luke to call upon more powerful vampiric abilities (having already been impressed by Luke's speed and reflexes) but is disappointed and disgusted to find Luke can't even heal his severed legs. Alucard then quips that in life (as a vampire), Luke was useless and pathetic. In response, Luke calls Alucard "a dog for the church of England," before the latter interrupts him. Luke later became "dog food" for Alucard's hell-hound. Alucard then mocks the demise of Luke Valentine, saying that the latter is nothing more than "dog shit."
Luke would later reappear from the hell-hound's corpse with only his upper body free. Confused with this situation, he is quickly killed again by Walter's strings and his and half of Baskerville's body are forced to attack Alucard. This manages to push Alucard for awhile but he eventually shoots Luke and stops the attack. As Walter goes to stab an severely injured Alucard in the heart, it is revealed to be Luke's body acting as a double. This allows Alucard (in his Girlycard form) to sneak up on him. Alucard then happily remarks at how surprised he is at how useful Luke was.
- Enhanced Speed/Reflexes: While his brother Jan, with the over-the-top strength and agility, is more of a basic vampire, Luke in particular demonstrates an extreme form of celerity. He can easily dodge bullets and slice his enemies to pieces before they could even react.
- Enhanced Durability: In the manga, he apparently took a point-blank shot to the chest from the Casull without visible damage. This is in contrast with the OVA, where he was shot in the head and regenerated the damage.
- Regeneration: Luke was capable of regenerating wounds such as being stabbed in the chest by spears and being shot in the head by the Casull. His regeneration isn't unlimited, however. When Alucard shot off his legs, he couldn't heal his wounds, and fell short on their battle.
- Sharpshooting Skills: Luke is skilled in sharpshooting. Going toe-to-toe with Alucard.
When Luke meets Alucard in the sub-levels, Luke seems to be quite condescending to Alucard in the beginning, but frightened towards the end where Alucard kills him. Luke claims that he will prove that immortality is a myth, once he has Alucard's head on a stick in the Hellsing OVA II. After Alucard tore off Luke's legs, Luke refers to Alucard as the dog of the Church of England, to which Alucard replies by calling Luke 'dog food", followed by devouring him as Baskerville. Alucard goes back to that comment while belittling what is left of Luke. "As a vampire you were just a pathetic piece of shit. Now you're nothing but dog shit."
Given how short their time on-screen together was, it's hard to say exactly how well the two got along. Obviously, Jan is very comfortable talking with Luke, chatting the entire way to the Hellsing Mansion. Luke, however, didn't seem too pleased with the conversation. Luke seems somewhat short with Jan, usually telling him to shut up when he starts rambling.
- "Please, be quiet."
— Luke while he's killing Hellsing personnel
- "Well, that's enough of that. Time to see what you can really do, Alucard."
- "It is a pleasure to make your acquaintance, Alucard. My name is Luke Valentine. I'm a real fan of your work."
- "Your name gets thrown around quite a bit, Alucard. The things people say...They tell stories of your amazing strength. How you glide through oceans of blood. They call you nightwalker. Beyond human. A monster whose powers radiate with the darkness that casts the shadow on darkness itself. Finally I get to know once and for all if there's anything more to you than rumor. When I was human I feared you. The stories, anyway. And when I gave up being human, when I became this, I began to admire you. You, the first great pioneer of immortality. No matter what the rest of us do really were all just chasing your shadow."
- "Yes, I know. And once I have your head on a stick, everyone else will know it too. So now I get to kill you and make my wildest dreams come true."
— Luke responding to Alucard claiming immortality is a myth
- "You've never seen anything like me before! I was designed to surpass you in every way! I was born to be your death!"
- "I told you, I'm completely different than any creature you've faced before! I have the sum total of your vampiric power at my disposal! All of it, and more!"
- "He's not even trying to dodge the bullets! Does he think he's indestructible?! Everything has a breaking point. He may act like he's not injured, but he's taking far more damage than I am! I can beat him!"
— Luke while shooting Alucard
- "What are you? WHAT IN THE HELL ARE YOU?!"
— Luke upon witnessing how powerful Alucard really is
- "Shut up!! You're nothing but the Hellsing family toy! A dog for the Church of England not even fit to call himself a vampire! A..."
- "What is this? What's happened to me, the hell's going on here?"
— After just emerging from the Baskerville's corpse
- Despite having a severed arm during his final [OVA II] moments, it is worth noting that he could regenerate inside Baskerville (with the exception of his legs).
- In OVA II, Alucard remarks near the end of their battle that he considers Luke "above even a Class A vampire"; in the anime, however, he simply saw him as a "Class A Vampire".
- It is worth noting that Luke Valentine is the only member of the Millennium Group to be killed by Alucard exactly the same way in every incarnation of the series (manga, Gonzo series and OVAs).
- The 9th Volume and OVA reveal that Luke, unlike his brother, holds a military rank within Millennium, namely Warrant Officer (the same as Schrödinger).
- It is interesting to note that the nationalities of the brothers is never made clear, although it seems they are from the United States, judging by their dialogue in the manga, which is devoid of British phrases such as "wanker" (which at least Jan would've used at some point), or German pronunciations (like "ve" instead of "we"), or, if they were thought to be British, the fact that they aren't at all phased that one of Millennium's plans is to wipe out London in what can be thought of as a terrorist attack.
- Despite the fact that in the manga, only Jan's hat is present in the Valentine brothers' shrine, in the 10th OVA, Luke's curved knife is now his shrine.
- Luke's gun is based off of a sawed off M1 Garand. There is an actual gun design called the "Enforcer" M1. They were produced mainly in the 60's and 70's by a variety of manufacturers, including Iver Johnson and Universal.