In October 1956, Washington University researcher Dr. William "Bill" Masters proposes a controversial study of human sexuality but is rejected by his university. Undaunted, he begins the study anyway and hires Virginia Johnson, a nightclub singer-turned-secretary, as his assistant.
Virginia has trouble telling Bill that she cannot accept his proposal, but Bill informs her that Scully has cancelled the study because he found out they were monitoring couples having sex. Bill believes it was Ethan who told Scully and fires Virginia on the spot for sleeping with him.
Bill and Virginia continue working at the brothel but Bill grows increasingly frustrated with the conditions. Betty recruits some men for the study but Bill is nonplussed to learn they are homosexual prostitutes.
With the study back in the hospital, Bill and Virginia recruit new participants on campus. Virginia has a one-night tryst with her ex-husband, George, who appears at the hospital, having signed up as a study participant.
Bill experiences anxiety about becoming a father. Ethan is worried about dating Barton Scully's daughter, Vivian, but she reassures him that they were meant to be together. Libby suffers a miscarriage.
In the aftermath of the miscarriage, Libby and Bill vacation in Miami, but Masters is drawn back into his work by a sexually adventurous couple next door. Virginia enlists Jane in an effort to debunk Freud's theory that a vaginal orgasm is more mature one than a clitoral one.
Virginia tries to find a new secretary. Bill tries to balance the study and his affair with his home life. Austin continues his sexual relationship with Margaret Scully, while Barton Scully is stabbed by a trio of gay-bashing thugs.
Bill and Virginia decide that the best way to record their study findings is to film them, but to do so they have to hire a cameraman. Johnson redoubles her efforts to get a college degree, but when she learns that Dr. DePaul's Basic Anatomy class is a requirement, DePaul refuses to give her class credit for her work with Masters.
Bill and Virginia decide to film external bodily reactions in the study, and hire a filmmaker named Lester to film Jane masturbating. Vivian wants a Christian wedding, and after Ethan reveals he's Jewish, he agrees to convert to Christianity.
A young woman named Flora Banks tells Bill and Virginia that she got pregnant from her participation in the study, and wants to know the identity of the father. Bill and Virginia disagree as to whether to maintain study confidentiality or divulge the father’s identity to Flora.
With Virginia no longer a part of his office, Bill accepts Libby's offer to temporarily serve as his secretary, in order to help him help compile the study work for a hospital-wide presentation. However, in the course of her work, she learns of a file on a couple who had sex in the study 23 times.
Bill gives the presentation on his and Virginia's research to the hospital, but it does not get the response he had hoped. Jane is shocked to see that the footage of her that she thought was destroyed is used in the presentation.
The presentation continues to affect the reputations of both Bill and Virginia; Bill is urged to get a new job after being fired from Maternity at the end of season one; Bill and Virginia confront their feelings towards each other.
Bill cuts sexual ties with Virginia after finding out that she was having relationships with other people, and a surprise visit by his mother, who offers to give him money to pay for his new clinic, doesn't help.
Virginia sees a psychiatrist while pretending to be one of their own subjects, and begins to worry about the effect her and Bill's relationship will have on Libby; Bill is bothered by his brother; Libby prepares to testify.
Bill admits his impotence and begs for Virginia's help resolving it; Virginia is intrigued when a man from CBS says he wants to air a special on it; Libby continues working for CORE; Lester and Barbara connect.
A television crew interviews Bill and Virginia, but Bill is repeatedly bothered by the censorship; George wants to take the kids to Europe for several weeks, to Virginia's dismay; Libby and Robert finally confront their feelings.
Virginia is caught off guard when she gets letters from her son serving in the war, and thinks he might be sick, which may keep her home as Bill and Virginia are urged to tour the country in order to increase book sales.
Bill is invited back to Washington University to deliver a speech after the success of the book; Virginia is surprised by the arrival of her parents; Margaret tries to adapt to a three-person relationship.
Bill and Virginia struggle to continue their relationship as Logan continues to pursue her; Bill's problems with his son escalate when Bill makes Johnny sign up for football; Libby discovers Joy's secret apartment; Virginia's mother continues to stir up trouble.
Bill reluctantly agrees to help resolve the sexual dysfunction of a gorilla just as he and Virginia are being interviewed by Newsweek; Virginia and Dan Logan continue seeing each other; Libby apologizes to Paul for revealing Joy's true feelings; Betty and Helen, after deciding they want a child, begin looking for sperm donors.
Virginia and Logan drop everything to relax in Las Vegas, but Virginia can't seem to put her work behind her; Bill starts his surrogacy program despite Virginia's prior skepticism; Libby is forced to confront memories of her past lover; Betty and Helen enlist Austin to help them have a baby.
Bill becomes desperate to reconnect with Virginia as she raises concerns about Bill's new surrogacy program; Dan plans on leaving Missouri; Libby tries to set Paul up with one of her friends; Austin loses custody of his kids and ends up staying with Betty and Helen.
Virginia's absence in Bill's love life continues to have an upsetting effect on him, and he ends up having to sacrifice his surrogacy program; Libby and Paul take their relationship to the next level; Tessa is caught in the act with her boyfriend.
While in New York, Bill orchestrates a dinner with Virginia, Dan, and Dan's wife, in hopes he will finally be reunited with Virginia; Libby and Paul's night together is interrupted when a detective shows up to investigate an accusation about Bill.
With their publisher eager to move forward with their new book and the police investigating Bill's recent indiscretions, all of Bill and Virginia's work is compromised as they are stabbed in the back by someone close to them; Virginia finally chooses between Dan and Bill.
In the wake of their fallout, Masters and Johnson search for purpose in a world without one another and look to work as their personal lives crash down around them. Libby finds herself alone for the first time since her marriage and discovers some unexpected allies.
Masters and Johnson turn to new partners in an effort to put the past behind them and focus on their expanding clinic. Given their fractured triangle, Libby reevaluates her relationship with Bill and Virginia.
Masters and Johnson adjust to new partners, but their respective cases have them second-guessing the instinct to remain separated. A romantic prospect pushes Libby to embrace life as a newly single woman of the sexual revolution.
Bill, Virginia, Libby, Betty, Lester, and Keller all end up at a swingers party hosted by Art and Nancy; Bill and Libby re-examine their complicated relationship; Virginia confronts her feelings toward Bill.
Virginia deals with insubordination in the clinic, and continues to try to reunite with Bill, despite his pulling away; Helen goes into labor and Betty is left dealing with the aftermath of her in-laws abrupt exit and an unforeseen tragedy.
Bill contacts a lover from his past, while he and Art try to help a possibly bisexual client continue with his marriage; Virginia and Nancy discover their clinic is under scrutiny from competing businesses; Louise from AA surprises Bill, in need of a favor.