Cervical cancer is a significant cause of female mortality worldwide, and early detection through regular screening is crucial for reducing mortality rates. However, in developing countries, the uptake of Pap smear tests (PST) is low, mostly due to cultural and social factors and a lack of knowledge. This cross-sectional study assessed knowledge, attitude, and practice of cervical cancer screening among practitioners working at primary healthcare centers in Saudi Arabia. Furthermore, the study aimed to identify the potential barriers that prevent female physicians from performing cervical cancer screening tests. A self-administrated, well-structured questionnaire was used to survey 95 female physicians, including residents, specialists, and consultants in several primary health care centers (PHCCs) in Jeddah managed by the Ministry of Health during September 2019. The results showed that 80% of participants knew about cervical cancer, and 97.8% were aware that PST is a screening tool. However, only 47% advised female patients to get tested for cervical cancer. The factors identified as barriers to test uptake included asymptomatic females (34%), lack of time on the part of the practitioner (24%), and a lack of evidence for risk factors (23%). Additionally, only 42.2% of the participating physicians had undergone a Pap smear test themselves. The study highlights the need for further research to assess HPV status in the population and explore the correlation between circumcision and cervical cancer, as well as polygamy and cervical cancer. The findings suggest that while a good level of knowledge about cervical cancer exists, there is a need to improve compliance with cervical cancer screening guidelines among female physicians in Saudi Arabia.