The original series was based on Professor Helen Witwell, a forensic pathologist based in Sheffield, whom McCrery had known while serving as a police officer. The programme originally followed the activities of a female pathologist, Professor Sam Ryan, played byAmanda Burton. However, Ryan's character departed early in the eighth series in 2004. There had been a succession of regular supporting characters, changing almost every series, but Dr Leo Dalton (William Gaminara) and Dr Harry Cunningham (Tom Ward), who were introduced in the sixth series in 2002, remained in the series and continued as lead characters following Ryan's departure, with Dalton replacing her as professor. A new character, Dr Nikki Alexander (Emilia Fox), was introduced to the team in the eighth series in 2004. While working as a forensic anthropologist, she appropriates facilities and software in the pathology department to analyse an Iron Age find, with the belated, bemused and begrudging approval of Dalton. Dr Alexander is able to assist in a set of cases being investigated by the team, as it turns out she had "worked in forensic pathology in Johannesburg for six months" and is Home Office certified to practise. She is of such assistance that she overcomes Leo's reluctance and, with Harry's support, is offered and accepts a position on the team.

Some early series were set in Cambridge, however, the show has since relocated toLondon, and more recently, Manchester. The programme is typically made up of a series of two-part stories, usually with six to eight episodes per series. The theme music for Silent Witness is a song called "Silencium" by John Harle. The incidental music for Silent Witness is written by BAFTA nominated composer Sheridan Tongue. In 1998, writer John Milne received an Edgar Award from the Mystery Writers of America for the second series episode "Blood, Sweat, and Tears". In the United States, the show aired during 'Mystery Monday' on BBC America. It airs in Norway using the name "Tause vitner" on the Norwegian Broadcast Channel NRK, in Sweden using the name "Tyst vittne". Both the Norwegian and Swedish titles are direct translations. The show is also broadcast on Swedish commercial channel Sjuan, in the Netherlands by public broadcaster KRO, Belgium on VRT channel Canvas and in Finland on the Finnish Broadcasting Company YLE using the name "Hiljainen todistaja", which is a direct translation of the original title.

Silent Witness has been criticised for gruesome and harrowing scenes as well as for its failure to convey the work of forensic pathologists accurately.[3] Nevertheless, fifteen years after its inception, it continues to achieve good audience ratings.[4] The approach of the show, portraying a pathologist as having an active role in the crime investigation, was parodied by British comedic duo French and Saunders as "Witless Silence". Dead Ringers also parodied Silent Witness, with Sam Ryan as an overconfident pathologist who makes incredibly specific guesses about the body; for example, "Just by looking I can tell that this was a man aged 35–37 called John, having an affair with his secretary", only to be proved wrong by one of her assistants, "No that's an onion bagel, I got for your lunch". She then refuses to accept her mistake claiming "Wrong? Oh, I'm never wrong. I'm forensics professor Sam Ryan PhD."

Although the show heavily focuses on areas of pathology, the police also play a heavy presence in each case. During latter series of the show, detectives and investigators tend to differ from episode to episode, with guest artists appearing in these roles. However, during the early years of the show, several characters appeared regularly to investigate each case.