Since 1998 in Japan, guidelines for cancer screening programs have been developed and revised by a research group funded by the Ministry of Health, Labour and Welfare. However, little is known about health professionals' awareness of and adherence to the cancer screening guidelines. Surveys were conducted by mailing questionnaires to two target groups of health professionals: local government officers of municipal cancer screening programs of 3327 municipalities in 47 prefectures (local government officers group; n = 3327) and councilors of an academic society dealing with a mass survey of gastroenterological cancer (expert group; n = 195). The questionnaire contained questions dealing with: (1) awareness of and adherence to the cancer screening guidelines published in 2001, and (2) basic knowledge of and attitude towards cancer screening. We compared the responses of the two groups. The response rate in both groups was approximately 65%. Over 70% of the respondents were aware of the cancer screening guidelines. However, 20% of the local government officers and 35% of the experts thought that non-recommended methods could be used for population-based screening. Fifty-six percent of the local government officers and 76% of the experts responded that there was no problem with using non-recommended methods for opportunistic screening. Almost all health professionals believed that screening was 'almost always a good idea'. Although the two groups' backgrounds differed, both did not sufficiently understand the evidence-based approach for cancer screening. To properly conduct evidence-based cancer screening, it is necessary that health professionals have an appropriate understanding of the guidelines.