They claim the Church has launched a hostile takeover bid for the Blue Coat school, Liverpool, which was founded in 1708 and has more Catholic than Anglican pupils.
The trustees say they will fight the move in the courts if necessary.
The diocese of Liverpool has asked the Government to rule on whether the grammar should become an Anglican school and is confident of its case. It says it has a legal responsibility to act following a tip-off that the school was wrongly designated as secular after Labour stripped it of its grant-maintained status in 1998.
The Church argues that, because documents published by the Charity Commission as recently as 2000 state that Anglican religious education must be given in Blue Coat, it is a C of E school in all but name.
But Sandy Tittershill, head of Blue Coat, said: "I have been working at this school for 40 years and I have never known it to be a church school.
We hold broadly Christian assemblies but have always been non-denominational, multi-cultural and mixed in every possible way.
"I don't believe there is any desire on behalf of the parents' body, the students or the staff to become a C of E school."
The school's roll of 900 includes Catholic, Jewish, Muslim, Hindu and Chinese pupils and he estimates that only between 20 and 30 per cent have Anglican backgrounds.
The diocese denies governors' suspicions that its real motive is to control assets worth pound;4.44 million belonging to the school's charitable foundation.
Jon Richardson, diocesan education director, said the foundation would retain control over the assets and its power to appoint a majority of governors.
"That is a matter of charity law that we could not change even if we wanted to," he said.
The Blue Coat governors won their last major battle in the mid- 1980s when Liverpool's then Militant-controlled council tried to withdraw the school's funding.