The Palestinian parliament speaker, led in shackles into an Israeli military court, has been charged with membership in an outlawed organisation.
Abd al-Aziz Dweik becomes the most senior of three dozen Hamas officials rounded up by Israel to be indicted so far.
Dweik said on Tuesday that he does not recognise the court's authority.
"It is a political trial, and I don't recognise it," Dweik said. "I am an elected official."
Dweik was brought into the courtoom in the Ofer Prison Camp with his feet shackled.
He wore a brown pajama-style outfit, and his normally long beard was trimmed.
The military prosecutor charged Dweik with membership and activity in an outlawed organisation.
It was not immediately clear how many years he could face in prison.
The next hearing was set for August 31.
The prosecutor said that at one point, Dweik called Hamas's supreme leader, Khalid Mishaal, and asked for large sums of money.
However, that allegation was not part of the indictment.
In all, Israel has arrested 30 Hamas politicians and five cabinet ministers, including Nasser Shaer, the deputy prime minister, in recent weeks.
Dweik said Shaer became his cellmate on Monday.
Dweik's lawyers complained that he was being held in unsanitary conditions and that his cell was full of cockroaches.
Dweik was arrested earlier this month.
At one point, he was taken to a Jerusalem hospital after complaining of chest pains. His lawyers said he also has a kidney problem.