Empowering lay screeners, such as pre-school teachers, on vision screening is a cost-effective way to ensure larger populations of children can be screened. Although the validity of lay screeners in conducting vision screening were reported in several studies, none showed data concerning improvement of the level of knowledge among lay screeners after completing vision screening training, which could indicate the effectiveness of the training program. This study aimed to determine the level of knowledge of pre-school teachers before and after attending a training program. Sixty pre-school teachers from Tabika and Taska KEMAS were randomly selected. The Study Group (n = 30) was given theory and practical training on vision screening, whereas the Control Group (n = 30) was only given brief verbal instructions on how to conduct the screening. A theory test containing 15 questions related to the training modules were administered to both groups, before and after their training/briefing respectively. The findings showed that the level of knowledge among preschool teachers in the Study Group (73.24 ± 11.73%) was significantly higher than the Control Group (56.22 ± 13.11%) (p < 0.01). There was also a significant improvement in the level of knowledge among pre-school teachers in the Study Group after the training (p<0.001), whereas no improvement was noted among preschool teachers in the Control Group (p = 0.636). This study shows the importance of conducting training for pre-school teachers prior to their involvement in conducting vision screening in order to deliver an effective vision screening program to the preschoolers.