What was Big Willy Shakes going for?
Lady Macbeth is trying to be reassuring. The thing is, she's not really the reassuring type. In Lady Macbeth's mind, being a woman interferes with her evil plans. To her, femininity means compassion and kindness, while masculinity is synonymous with "direst cruelty." So she tries to stay away from the whole womanly bit. And that includes reassuring her husband.
She tells him they can't change the past ("what's done is done") and that he should just get over it. Too bad it's not that easy. Macbeth is already feeling guilty for murdering King Duncan.
We would also like to point out that Lady Macbeth eerily echoes this phrase in her final appearance in the play. By Act 5, Lady Macbeth has been saying and doing some freaky things on her nightly strolls. She sleepwalks, continuously trying to wash the imaginary blood from her hands. As she walks, she mutters, "what's done cannot be undone" (5.1.63-4).
We're thinking this is about more than actual blood staining her hands. In fact, we think blood is a symbol for guilt in this play. Lady Macbeth has grown so ill that the doctor says there's nothing he can do to help her. It's not a physical sickness a doctor can treat.
Her eerie confession to the murders shows she is not actually okay with them. She might have been the one pushing Macbeth to grow up and get over it in this first scene; but by the end of the play she is totally guilt ridden as well. It looks like it's a darn shame that her words are true. What is done is done. There are no take backs when it comes to murder.