Background: Colostrum is the first breast milk produced after birth and is important for promotion of health and prevention of infections of the newborn. Though breastfeeding practices are well known but the necessity of colostrum feeding is still poorly understood by mothers.Objective: To assess Knowledge, Attitude, Practices and associated factors towards colostrum feeding among mothers of infants in ambo district of west Shoa zone, Oromia, Ethiopia.Methods: A community-based cross-sectional study was conducted from March to April 2019 among 429 mothers of infants selected by simple random sampling technique. The data were collected using structured questionnaire, focus group discussion & key informant interview by semi structured guiding questionnaire. Descriptive analysis like frequency, percentage, & mean were performed. Binary and multiple logistic regression analysis were employed to identify associated factors. Variables with p-value <0.05 with 95% confidence interval identified statistically significant.Results: Among the respondents, 278 (64.8%) mothers had good knowledge & 250 (58%) had favorable attitude. But colostrum feeding was practiced only by 227(56.5%). Mothers of neonates without any illness/finding at birth (AOR =14.87 [ 95% CI: 5.00-44.27]) & postnatal care within the first 2-3 days (AOR = 3.48 [95% CI: 1.23-9.85]) were positively associated factors; but unwanted pregnancy/birth (AOR = 0.243 [95% CI: .113-.527]), mothers suffering from any health problems during pregnancy/birth (AOR = 0.380 [95% CI: .161-.903]), lack of counseling (AOR = 0.264 [95% CI: .103-.675]), home delivery (AOR = 0.239 [95% CI: .111-.516]), & lack of information on colostrum (AOR = 0.040 [95% CI: .013-.125]) were negatively associated with colostrum feeding practices. Conclusions: Majority of respondents had good knowledge & favorable attitude but with poor practices of colostrum feeding. Good neonatal health at/soon after birth & postnatal care attendance within two to three days were positively associated with colostrum feeding practices. But, unwanted pregnancy/birth, any sickness of mother at birth/during pregnancy, lack of counseling on colostrum feeding, home delivery, & lack of information on colostrum feeding were less likely to feed colostrum.Strengthening family planning service, antenatal care, institutional delivery, postnatal care, information provision for counseling & the revision of DHIS2 for inclusion of colostrum feeding indicators were recommended interventions.