Objectives: This study attempted to understand the relationship between handwashing knowledge, attitude and practice of middle school students and to provide the basic information for handwashing training of the students and help them to form a healthy habit. Methods: Data was collected by a standardized self-administered questionnaire between Sep. 20 to Oct. 6 2006 with a total of 710 students in 1, 2, 3 grades at 6 middle schools in metropolitan cities(490 students from four schools situated in an urban center, 220 from 2 in a suburban district). The data was then analyzed using the SPSS WIN 12.0 program, employing many statistical techniques such as chi-square($x^2$) test, t-test, ANOVA analysis with post hoc test, correlations analysis, and regression analysis. Results: The results of the study were as follows. First, total number of handwashing times of middle school students is 6.69 per day. 66.1% of the students used soap and most of the students washed hands for $6{sim}10$ seconds(44.7%). Second, in the knowledge of handwashing, "both of the parents"(p<0.01), "catholic", "city"(p<0.05) were the highest and the practice of handwashing was active with "city", "high economic level"(p<0.05) and "both of the parents"(p<0.01). In the experience of teaching handwashing, "Yes" was 24.3% which was statistically significant with handwashing knowledge(p<0.05), attitude and practice(p<0.01). Third, in the correlation of handwashing knowledge, attitude and practice, there was statistical significancy between handwashing attitude and knowledge(p<0.01) and showed positive correlation with the knowledge(.534). The practice of handwashing has meaningful difference from the knowledge and the attitude and the coefficient of correlation shows positive co-relation in knowledge(.335) and attitude(0525). Fourth, based on the result for regression analysis with handwashing knowledge, attitude and practice as dependent variables, handwashing knowledge showed statistical significancy with sex, type of school(p<0.05) and residential area(p<0.01). There were also statistical significancy between handwashing attitude and knowledge(p<0.01) and between handwashing practice and religion, knowledge(p<0.05) and economic level, attitude(p<0.01). Conclusion: There were consistent results with handwashing knowledge, attitude and practice. If they had better knowledge, their attitude was more positive and if they had better knowledge and more positive attitude, their practice was active. The knowledge, attitude and practice of the students who learned about handwashing were higher than those of the students who had no experience of learning handwashing. To enhance handwashing habit of middle school students, the handwashing environment should be maintained. The training plans should also be made according to sex, type of school, economic level, residential area, and the consistent study on handwashing training is required.