Organ transplantation is still the treatment of choice for end-stage organ failure. Despite the establishment of the Saudi Center for Organ Transplantation (SCOT), the organ supply still does not meet the increasing demand. This study aims to assess the knowledge of Saudis about organ donation both among the public and the medical staff and to assess the trend in the change from 1996 until now. A cross-sectional study was conducted among Saudis above the age of 18 years. A self-administered, 25-item, questionnaire was completed by participants including both outpatients and medical staff at King Abdulaziz Medical City using a convenient sampling technique. A total of 376 individuals answered the questionnaire; 238 (63.1%) heard about SCOT, of whom 124 (70.5%) were males. Health-care professionals had better attitude and knowledge on organ donation (P <0.05). Compared to the results from 1996to 2017, there was (51.22%) to (30.4%) decrease in people who allowed their families to donate their organs; on the other hand, there was an increase in the percentage of people who had the correct knowledge that Islam supports organ donation. Unsatisfactory levels of awareness impacted negatively on the knowledge and attitude of the population that is showed by the minor improvements over the years, and the slight proportion of participants who owned a donor card. Although 97.6% of the participants knew about the concept of organ donation, only 66.5% were familiar with the positive Islamic opinion. Based on the results of the comparison with the study in 1996, we conclude that more efforts from both the governmental and religious authorities are needed to increase awareness.