- Olga Sergeyevna Prozorova (Olga) – The eldest of the three sisters, she is the matriarchal figure of the Prozorov family though at the beginning of the play she is only 28 years old. Olga is a teacher at the high school, where she frequently fills in for the headmistress whenever the latter is absent. Olga is a spinster and at one point tells Irina that she would have married "any man, even an old man if he had asked" her. Olga is very motherly even to the elderly servants, keeping on the elderly nurse/retainer Anfisa, long after she has ceased to be useful. When Olga reluctantly takes the role of headmistress permanently, she takes Anfisa with her to escape the clutches of the heartless Natasha.
- Maria Sergeyevna Kulygina (Masha) – The middle sister, she is 23 at the beginning of the play. She married her husband, Kulygin, when she was 18 and just out of school. When the play opens she has been disappointed in the marriage and falls completely in love with the idealistic Lieutenant-Colonel Vershinin. They begin a clandestine affair. When he is transferred away, she is crushed, but returns to life with her husband, who accepts her back despite knowing what she has done. She has a short temper, which is seen frequently throughout the play, and is the sister who disapproves the most of Natasha. Onstage, her directness often serves as a tonic to the melodrama, and her wit comes across as heroic. Her vitality provides most of the play's surprisingly plentiful humour. She was trained as a concert pianist.
- Irina Sergeyevna Prozorova – The youngest sister, she is 20 at the beginning of the play. It is her "name day" at the beginning of the play and though she insists she is grown-up she is still enchanted by things such as a spinning top brought to her by Fedotik. Her only desire is to go back to Moscow, which they left eleven years before the play begins. She believes she will find her true love in Moscow, but when it becomes clear that they are not going to Moscow, she agrees to marry the Baron Tuzenbach, whom she admires but does not love. She gets her teaching degree and plans to leave with the Baron, but he is shot by Solyony in a pointless duel. She decides to leave anyway and dedicate her life to work and service.
- Andrei Sergeyevich Prozorov (Andrey) – The brother of the three sisters. In Act I, he is a young man on the fast track to being a Professor in Moscow. In Act II, Andrei still longs for his old days as a bachelor dreaming of life in Moscow, but is now, due to his ill-conceived wedding to Natasha, stuck in a provincial town with a baby and a job as secretary to the County Council. In Act III, his debts have grown to 35,000 rubles and he is forced to mortgage the house, but does not tell his sisters or give them any shares in the family home. Act IV finds Andrei a pathetic shell of his former self, now the father of two. He acknowledges he is a failure and laughed at in town for being a member of the village council whose president, Protopopov, is cuckolding him.
- Natalia Ivanovna (Natasha) – Andrei's love interest at the start of the play, later his wife. She begins the play as an awkward young woman who dresses poorly and hides her true nature. Much fun is made of her ill-becoming green sash by the sisters, and she bursts into tears. She apparently has no family of her own and the reader never learns her maiden name. Act II finds a very different Natasha. She has grown bossy and uses her relationship with Andrei as a way of manipulating the sisters into doing what she wants. She has begun an affair with Protopopov, the head of the local council (who is never seen), and cuckolds Andrei almost flagrantly. In Act III, she has become even more controlling, confronting Olga head on about keeping on Anfisa, the elderly, loyal retainer, whom she orders to stand in her presence, and throwing temper tantrums when she doesn't get her way. Act IV finds that she has inherited control of the house from her weak, vacillating husband, leaving the sisters dependent on her, and planning to radically change the grounds to her liking. It is arguably the vicious, manipulative, self-absorbed Natasha, who cares for no one besides her own children, Bobik and Sofia, upon whom she dotes fatuously, who is the complete victor by the end of the play.
- Fyodor Ilyich Kulygin – Masha's older husband and the Latin teacher at the high school. Kulygin is a jovial, kindly man, who truly loves his wife, and her sisters, although he is very much aware of her infidelity. In the first act he seems almost foolish, giving Irina a gift he has already given her, and joking around with the doctor to make fun of Natasha, but begins to grow more and more sympathetic as Masha's affair progresses. During the fire in Act 3, he confesses to Olga that he might have married her – the fact that the two would probably be very happy together is hinted at many times throughout the show. Throughout the show, often at the most serious moments, he often tries to make the other characters laugh in order to relieve tension, and while that doesn't always work, he is able to give his wife comfort through humor in her darkest hour at the show's climax. At the end of the play, although knowing what Masha had been doing, he takes her back and accepts her failings.
- Aleksandr Ignatyevich Vershinin – Lieutenant-colonel commanding the artillery battery, Vershinin is a true philosopher. He knew the girls' father in Moscow and they talk about how when they were little they called him the "Lovesick Major". In the course of the play, despite being married, he enters into an affair with Masha but must end it when the battery is transferred. He frequently mentions how his wife regularly attempts suicide (and he has two daughters), but he seems to have become inured to his domestic suffering. His first act speech about the hope he has for civilization speaks directly to Masha's melancholic heart, and, upon hearing it, she declares "I'm staying for lunch."
- Baron Nikolaj Lvovich Tuzenbach – A lieutenant in the army and not deemed handsome, Tuzenbach often tries to impress Irina, whom he has loved for five years. He quits the Army to go to work in an attempt to impress her. He is repeatedly taunted by Solyony and between Acts III and IV, he retaliates and prompts Solyony to declare a duel. He is killed in the duel, thus his and Irina's union is forlorn.
- Staff Captain Vassily Vasilyevich Solyony – A captain in the army, Solyony is a social misfit and a rather modern type of anti-hero. He is in love with Irina and tries to put down the Baron to make himself look better, but Irina finds him crude and unappealing. He spends much of his time mocking the Baron, who is the closest thing he has to a friend, and ends up killing him in a pointless duel. He is said to have a remarkable resemblance to the poet Lermontov in both face and personality, often quoting him. He always carries a small perfume bottle which he frequently (almost pathologically) sprinkles his hands and body with; it is later revealed that he does it to mask the smell of corpses on him.
- Ivan Romanovich Chebutykin – Sixty years old and an army doctor, Chebutykin starts off as a fun, eccentric old man who exults in his place as family friend and lavishes upon Irina the expensive gift of a samovar. Later on in Act III, while drunk, he suffers an existential crisis and reveals to all about Natasha's and Protopopov's affair. In Act IV however, he seems to have come to terms with his crisis or perhaps been broken by it. He loved the mother of the sisters (whose name is never mentioned) but she was married.
- Aleksej Petrovich Fedotik – A sub-lieutenant, Fedotik hangs around the house and tries to express his love to Irina by buying her many gifts. He also is an amateur photographer, and takes photos of the group and Irina. In Act III, he loses all his belongings in the fire, but retains his cheerful nature.
- Vladimir Karlovich Rode – Another sub-lieutenant, Rode is a drill coach at the high school.
- Ferapont – Door-keeper at the local council offices, Ferapont is an old man with a partial hearing loss. He repeatedly blurts out random facts, usually relating to Moscow.
- Anfisa – An elderly family retainer and former nurse, Anfisa is 81 years old and has worked forever for the Prozorov family. Natasha begins to despise her for her feebleness and threatens to throw her out but Olga rescues her, taking her to live at Olga's teacher's flat.
The play has several important characters who are talked about frequently, but never seen onstage. These include Protopopov, head of the local Council (albeit unseen on stage) and Natasha's lover; Vershinin's suicidal wife and two daughters; and Andrey and Natasha's children Bóbik and Sofia. JL Styan contends in his The Elements of Drama that in the last act Chekhov revised the text to show that Protopopov is the real father of Sofia: "The children are to be tended by their respective fathers" — Andrey pushes Bóbik in his pram, and Protopopov sits with Sofia.