Background: College students form a large, healthy, and active population of potential blood donors; their recruitment and retention would immensely help meet the demand of safe blood. The knowledge and practice of blood donation among students are reportedly diverse. In Nepal, medical students donate more and are more knowledgeable than non-medical, students but there could be differences between medical students of different colleges. Methods: A cross-sectional comparative study and using self-administered questionnaire was conducted in the students of two randomly chosen private medical colleges of Nepal – Nepal Medical College (NMC) of Kathmandu and Chitwan Medical College (CMC) of Chitwan. Results: Total 279 students, 106 in CMC and 173 in NMC, participated. Students of CMC were older (mean age 21.16±1.353 vs 19.32±1.094 of NMC), more knowledgeable (average knowledge score 55.2% vs 52.6% of NMC), donated more (50.9% donated vs 25.4% of NMC), participated more in organizing blood camps (37.7% vs 19.1% of NMC), and also donated more often. More boys donated; CMC consisted of 60% boys, the sex ratio reversed in NMC. Donation practice had significant correlations with gender (male propensity) and older age but not with knowledge. Conclusion: Significant differences exist in the knowledge and practice of blood donation among students of different medical colleges. Organizing blood donation camps routinely in colleges would encourage more students; especially younger, girls, and those having difficulty with time and information.