Who were the only possible witnesses of the murder in Macbeth?
There were five people directly involved in the murder scene: Macbeth, Lady Macbeth, Duncan, and Duncan's two servant-guards.
Macbeth commits the murder alone, after his wife drugs the guards into unconsciousness, and Duncan is asleep, so technically no one but Macbeth is actually awake enough to witness the murder itself. Lady Macbeth later goes to the room herself to cover the guards in blood, to pass suspicion onto them. Macbeth ends up killing the guards anyway, as a means of ensuring that their innocence can never be revealed.
Thus, the only living witnesses are Macbeth and his wife, which secures their alibi, even if it doesn't dispel the suspicions surrounding them. Additionally, they are both plagued by the sense that the blood from the murder was a "witness" to their crimes, and Lady Macbeth's sleepwalking and eventual suicide indicate that their own subconscious minds, or their consciences, were acting as witnesses to the murder.