The state's Department of Health has issued an order requiring all state workers, health care workers, and school teachers to be vaccinated or get weekly COVID-19 testing.
Local health care workers and school district staff lined the sidewalk near Fresno State.
"Our freedoms are being taken away right before our eyes and we have the right to choose," said Fresno Unified employee Veronica Vance.
California's vaccine mandate comes as state officials aim to fight rising coronavirus infections, mostly among the unvaccinated.
Doctors, including top U.S. infectious disease expert Anthony Fauci, say the vaccines are 'safe and effective in preventing hospitalizations and deaths from COVID-19.'
"When you work in a setting that's very high-risk like a healthcare setting, or a congregate setting, you work with people who can get really really sick with this disease. So that's why I think it's your responsibility to get vaccinated - not to protect yourself only but also the people that you're working with so that you don't transmit the disease to them," said UCSF Fresno Medicine Physician Dr. Sukhjit Dhillon.
But protestors are not convinced.
"I will not get the vaccine, I'm 100% sure of that. It's not FDA-approved," said Vance.
Vance along with others who don't plan on getting the COVID-19 vaccine said they're preparing to lose their jobs.
"They're not going to just quit their jobs but if they have to be terminated that's what's going to happen," she said.