Romero drew inspiration from Richard Matheson's I Am Legend (1954), a horror novel about a plague that ravages a Futuristic Los Angeles. The infected in I Am Legend become vampire-like creatures and prey on the uninfected. Discussing the creation of Night of the Living Dead, Romero remarked, "I had written a short story, which I basically had ripped off from a Richard Matheson novel called I Am Legend." Romero further explained:

George Cameron Romero, the son of Director George A. Romero, has developed Rise of the Living Dead, a prelude to his father's classic pitched with the working title Origins. The film tracks a six-year period leading up to the story told by his father. George Cameron Romero's script is intended to be an homage to his father's work, a terrifying glimpse into the political hot bed that was the mid-to-late 1960s and a bookend piece to his father's original story. Despite raising funds for the film on Indiegogo in 2014, the film has yet to go into production as of November 2017.

The opening title music with the car on the road had been used in a 1961 episode of the TV series Ben Casey entitled "I Remember a Lemon Tree" (that piece of music accompanying each time that George C. Scott's character, a Doctor who is secretly a drug addict, is injecting himself with morphine), and is also featured in an episode of Naked City entitled "Bullets Cost Too Much". Most of the music in the film had previously been used on the Soundtrack for the science-fiction B-movie Teenagers from Outer Space (1959). The eerie musical piece during the tense scene in the film where Ben finds the rifle in the closet inside the farmhouse as the radio reports of mayhem play in the background, can be heard in longer and more complete form during the opening credits and the beginning of The Devil's Messenger (1961) starring Lon Chaney, Jr. Another piece, accompanying Barbra's FLIGHT from the cemetery zombie, was taken from the score for The Hideous Sun Demon (1959). According to WRS, "We chose a selection of music for each of the various scenes and then George made the final selections. We then took those selections and augmented them electronically". Sound tech R. Lococo's choices worked well, as film Historian Sumiko Higashi believes that the music "signifies the nature of events that await."

Official film adaptations of Matheson's novel appeared in 1964 as The Last Man on Earth, in 1971 as The Omega Man, and the 2007 release I Am Legend. Matheson was not impressed by Romero's interpretation, feeling that "It was ... kind of cornball", though he later said, "George Romero's a nice guy, though. I don't harbor any animosity toward him."

Props and special effects were fairly simple and limited by the budget. The blood, for Example, was Bosco Chocolate Syrup drizzled over cast members' bodies. Consumed flesh consisted of roasted ham and entrails donated by one of the actors, who also owned a chain of butcher shops. Costumes consisted of second-hand clothing from cast members and Goodwill. Zombie makeup varied during the film. Initially makeup was limited to white skin with blackened eyes; but as filming progressed mortician's wax was used to simulate wounds and decaying flesh. As filming was not linear, the piebald faces appear sporadically. Eastman supervised the special effects, wardrobe and makeup. Filming took place between June and December 1967 under the working title Night of Anubis and later Night of the Flesh Eaters. The small budget led Romero to shoot on 35 mm black-and-white film. The completed film ultimately benefited from the decision, as film Historian Joseph Maddrey describes the black-and-white filming as "guerrilla-style", resembling "the unflinching authority of a wartime newsreel". Maddrey adds, it "seem[s] as much like a documentary on the loss of social stability as an exploitation film".

Night of the Living Dead entered the public domain in the United States because the original theatrical distributor, the Walter Reade Organization, neglected to place a copyright indication on the prints. In 1968, United States copyright law required a proper notice for a work to maintain a copyright. Image Ten displayed such a notice on the title frames of the film beneath the original title, Night of the Flesh Eaters. The distributor erroneously removed the statement when it changed the title.

Despite the controversy, five years after the premiere Paul McCullough of Take One observed that Night of the Living Dead was the "most profitable horror film ever [...] produced outside the walls of a major studio". The film had earned between $12 and $15 million at the American box office after a decade. It was translated into more than 25 languages and released across Europe, Canada and Australia. Night of the Living Dead grossed $30 million internationally, and the Wall Street Journal reported that it was the top-grossing film in Europe in 1969.

Night of the Living Dead was the first feature-length film directed by George A. Romero. His initial work involved filming shorts for Pittsburgh public broadcaster WQED's children's series Mister Rogers' Neighborhood. Romero's decision to direct Night of the Living Dead essentially launched his career as a horror Director. He took the helm of the sequels as well as Season of the Witch (1972), The Crazies (1973), Martin (1978), Creepshow (1982) and The Dark Half (1993). Critics saw the influence of the horror and science-fiction films of the 1950s in Romero's directorial style. Stephen Paul Miller, for instance, witnessed "a revival of fifties schlock shock... and the army general's television discussion of military operations in the film echoes the often inevitable calling-in of the army in fifties horror films". Miller admits that "Night of the Living Dead takes greater relish in mocking these military operations through the general's pompous demeanor" and the government's inability to source the zombie epidemic or protect the citizenry. Romero describes the mood he wished to establish: "The film opens with a situation that has already disintegrated to a point of little hope, and it moves progressively toward absolute despair and ultimate tragedy." According to film Historian Carl Royer, Romero "employs chiaroscuro (film noir style) lighting to emphasize humanity's nightmare alienation from itself."

Night of the Living Dead led to five subsequent films between 1978 and 2010, also directed by Romero, and inspired two remakes; the most well-known remake was released in 1990, directed by Tom Savini.

A Soundtrack album featuring music and dialogue cues from the film was compiled and released on LP by Varèse Sarabande in 1982; it has never been reissued on CD. In 2008, recording group 400 Lonely Things released the album Tonight of the Living Dead, "an instrumental album composed entirely of ambient music and sound effects sampled from Romero's 1968 horror classic".

The first remake, debuting in 1990, was directed by special effects Artist Tom Savini. It was based on the original screenplay, but included more gore and a revised plot that portrayed Barbra (Patricia Tallman) as a capable and active heroine. Tony Todd played the role of Ben. Film Historian Barry Grant saw the new Barbra as a corrective on the part of Romero. He suggests that the character was made stronger to rectify the depiction of female characters in the original film.

The film received a VHS release in 1993 through Tempe Video. In 1998, Russo's revised version of the film, Night of the Living Dead: 30th Anniversary Edition, was released on VHS and DVD by Anchor Bay Entertainment. In 2002, Elite Entertainment released a special edition DVD featuring the original cut. To celebrate the 40th anniversary of the film, Dimension Extreme released a restored print of the film on DVD.

In 1999, co-writer John A. Russo released a modified version called Night of the Living Dead: 30th Anniversary Edition. He filmed additional scenes and recorded a revised Soundtrack composed by Scott Vladimir Licina. In an interview with Fangoria magazine, Russo explained that he wanted to "give the movie a more modern pace." Russo took liberties with the original script. The additions are neither clearly identified nor even listed. However, Entertainment Weekly reported "no bad blood" between Russo and Romero. The magazine quoted Romero as saying, "I didn't want to touch Night of the Living Dead". Critics panned the revised film, notably Harry Knowles of Ain't It Cool News. Knowles promised to permanently ban anyone from his publication who offered positive criticism of the film.

The second remake was in 3-D and released in September 2006 under the title Night of the Living Dead 3D, directed by Jeff Broadstreet. Unlike Savini's 1990 film, Broadstreet's project was not affiliated with Romero. Broadstreet's film was followed in 2012 by the prequel Night of the Living Dead 3D: Re-Animation.

On September 15, 2009, it was announced that Simon West was producing a 3D animated retelling of the original movie, originally titled Night of the Living Dead: Origins 3D and later re-titled Night of the Living Dead: Darkest Dawn. The movie is written and directed by Zebediah de Soto. The voice cast includes Tony Todd as Ben, Danielle Harris as Barbra, Joseph Pilato as Harry Cooper, Alona Tal as Helen Cooper, Bill Moseley as Johnny, Tom Sizemore as Chief McClellan and newcomers Erin Braswell as Judy and Michael Diskint as Tom.

A collaborative animated project known as Night of the Living Dead: Reanimated was screened at several film festivals and was released onto DVD on July 27, 2010, by Wild Eye Releasing. This project aims to "reanimate" the 1968 film by replacing Romero's celluloid images with animation done in a wide variety of styles by artists from around the world, laid over the original audio from Romero's version. Night of the Living Dead: Reanimated premiered theatrically on October 10, 2009, in Ramsey, New Jersey at the Zombie Encounter and Film Festival. Night of the Living Dead: Reanimated was nominated in the category of Best Independent Production (film, documentary or short) for the 8th Annual Rondo Hatton Classic Horror Awards, but lost to American Scary, a documentary on television horror movie hosts.

Director Doug Schulze's 2011 film Mimesis: Night of the Living Dead relates the story of a group of horror film fans who become involved in a "real-life" version of the 1968 film.

Some reviewers cited the film as groundbreaking. Pauline Kael called the film "one of the most gruesomely terrifying movies ever made – and when you leave the theatre you may wish you could forget the whole horrible experience. . . . The film's grainy, banal seriousness works for it – gives it a crude realism". A Film Daily critic commented, "This is a pearl of a horror picture that exhibits all the earmarks of a sleeper." While Roger Ebert criticized the matinée screening, he admitted that he "admires the movie itself". Critic Rex Reed wrote, "If you want to see what turns a B movie into a classic [...] don't miss Night of the Living Dead. It is unthinkable for anyone seriously interested in horror movies not to see it."

Romero revolutionized the horror film genre with Night of the Living Dead; according to Almar Haflidason of the BBC, the film represented "a new dawn in horror film-making". The film has also effectively redefined the use of the term "zombie". While the word "zombie" itself is never used—the word used in the film is ghoul—Romero's film introduced the theme of zombies as reanimated, flesh-eating cannibals. Romero himself didn't initially consider the antagonists in the film zombies, later saying "I never thought of my guys as zombies, when I made the first film ... To me, zombies were still those boys in the Caribbean doing the wetwork for [Bela] Lugosi." The film and its successors spawned countless imitators, in cinema, television and video Gaming, which borrowed elements invented by Romero. Night of the Living Dead ushered in the splatter film subgenre. As one film Historian points out, horror prior to Romero's film had mostly involved rubber masks and costumes, cardboard sets, or mysterious figures lurking in the shadows. They were set in locations far removed from rural and suburban America. Romero revealed the power behind exploitation and setting horror in ordinary, unexceptional locations and offered a template for making an "effective and lucrative" film on a "minuscule budget". Slasher films of the 1970s and 80s such as John Carpenter's Halloween (1978), Sean S. Cunningham's Friday the 13th (1980), and Wes Craven's A Nightmare on Elm Street (1984) "owe much to the original Night of the Living Dead", according to author Barry Keith Grant.

On October 3, 2017, Mill Creek Entertainment released a standard 1080p version of the film on Blu-ray in the United States. This was followed by a 4K restoration Blu-ray released by The Criterion Collection on February 13, 2018, sourced from a print owned by the Museum of Modern Art and acquired by Janus Films. The new 4K restoration is protected by copyright. This release also features a workprint edit of the film under the title of Night of Anubis, in addition to various bonus materials.

Starting in 2015, and working from the original camera negatives and audio track elements, a 4K digital restoration of Night of the Living Dead was undertaken by the Museum of Modern Art (MoMA) and The Film Foundation. The fully restored version was shown at MoMA in November 2016 as part of To Save and Project: The 14th MoMA International Festival of Film Preservation. This same restoration was released on Blu-ray by Criterion Collection on February 13, 2018.