Less developed areas have the highest incidence rates of cervical cancer, which vary by geographic region. If the right lifestyle changes are made, all cervical cancer risk factors can be avoided. Furthermore, vaccines have been created to guard against a number of the high-risk strains of the human papillomavirus (HPV). Not all women are aware of cervical cancer or how to prevent it, even though it is preventable. Therefore, every woman must assess level of knowledge regarding HPV prevention and cervical cancer. We applied a validated questionnaire between January and June 2021 to the female health workers at Ondokuz Mayıs University Hospital to assess their knowledge of cervical cancer and HPV protection as well as their attitudes toward it. Our findings indicate that women's knowledge on this topic is lacking, especially regarding the most recent advancements in HPV testing and vaccination-based cervical cancer prevention. Only 99% of the women who responded to the survey were actually aware that HPV testing was available. Although the incidence of cervical cancer is low, this figure is subject to change, particularly in light of the economic crisis and rising immigration rates, which could lead to higher rates of HPV infection in the population if no action is taken. Although the HPV vaccine has been available for some time, the Turkish government does not recommend it as a routine vaccine as part of the National Immunization Program. Even among the highly educated workforce in the healthcare industry, there is a lack of understanding about HPV vaccine. Healthcare professionals should be fully informed about the HPV vaccine for their own benefit so that they can discuss it with their patients. This study aimed to remove vaccination barriers and assess healthcare professionals' current knowledge of cervical cancer and HPV vaccination in a tertiary hospital.