The painting represents the scene of The Last Supper from the final days of Jesus as narrated in the Gospel of John 13:21, when Jesus announces that one of his Twelve Apostles would betray him.

The Last Supper specifically portrays the reaction given by each apostle when Jesus said one of them would betray him. All twelve apostles have different reactions to the news, From left to right:

• Bartholomew, James, Son of Alphaeus and Andrew form a group of three, all are surprised.
• Judas, Peter and John form another group of three. Judas is wearing green and blue and is in shadow, looking rather withdrawn and taken aback by the sudden revelation of his plan. He is clutching a small bag, perhaps signifying the silver given to him as payment to betray Jesus, or perhaps a reference to his role within the 12 disciples as treasurer. He is the only person to have his elbow on the table; traditionally a sign of bad manners. Peter looks angry and is holding a knife pointed away from Christ, perhaps foreshadowing his violent reaction in Gethsemane during Jesus' arrest. The youngest apostle, John, appears to swoon.
• (Jesus)
• Thomas, James the Greater and Philip are the next group of three. Thomas is clearly upset; James the Greater looks stunned, with his arms in the air. Meanwhile, Philip appears to be requesting some explanation.
• Matthew, Jude Thaddeus and Simon the Zealot are the final group of three. Both Jude Thaddeus and Matthew are turned toward Simon, perhaps to find out if he has any answer to their initial questions.