The blaze, blamed on an electrical fault, hit the Abu Sifin church located in the densely populated Imbaba neighbourhood west of the Nile river, part of Giza governorate.
Witnesses described how people rushed into the burning house of worship to rescue those trapped but were soon overwhelmed by the heat and the deadly smoke.
"Everyone was carrying kids out of the building," Ahmed Reda Baioumy, who lives next to the church, told AFP.
"But the fire was getting bigger and you could only go in once or you would asphyxiate."
The Egyptian Coptic Church and the health ministry reported 41 dead and 14 injured in the blaze before emergency services said they had brought the blaze under control.
Egypt's President Abdel Fattah al-Sisi declared on his Facebook page in the morning: "I have mobilised all state services to ensure that all measures are taken."
He later said he had "presented his condolences by phone" to Coptic Pope Tawadros II, who has been the head of the denomination in Egypt since 2012.
The interior ministry later said that "forensic evidence revealed that the blaze broke out in an air-conditioning unit on the second floor of the church building".
Father Farid Fahmy, of another nearby church in Imbaba, told AFP the fire was caused by a short circuit.
"The power was out and they were using a generator," he said. "When the power came back, it caused an overload."