Introduction: Colostrum is yellow to orange colored milk produced during a first few days of lactation and it is rich in nutrients and antibodies that have great role in health of the new born baby. Despite the world health organization recommends that every new born baby has to feed breast milk with colostrum within one hour of delivery; poor colostrum feeding practice is still a common problem in Ethiopia. Objective: This study aims to evaluate practice of colostrum feeding and associated factors among mothers who attended child immunization clinic in governmental health facilities of Dire Dawa administrative city, Eastern Ethiopia, 2021 Method: Institution based cross-sectional study was employed from February 1–30/2021 in governmental health facilities of Dire Dawa administrative city among 292 mothers with baby less than or equal to 12 months. The data were collected by systematic random sampling technique and analysed using Statistical Package of Social Sciences 25.0 version. Bivariate and multivariate logistic regression analyses was employed to estimate the crude and adjusted odds ratio with confidence interval of 95% and P value of less than 0.05 considered statistically significant. Result: In this study, 68.8% with [95% CI: 63.4-74.3] of the mothers had good practice of colostrum feeding. Living in urban [AOR=2.52, 95% CI=1.08-5.88],secondary and above education [AOR=2.79, 95% CI=1.12-6.98], having history of antenatal visit [AOR=3.12, 95% CI=1.12-9.21], counselled on timely initiation of breast feeding [AOR=2.59, 95%, CI=1.02-6.59], counselled on colostrum feeding [AOR=2.65, 95%, CI=1.12-6.30], birth attended by health professional[AOR=3.20, 95% CI=1.23-8.31], getting information from health professional AOR=3.89, 95%, CI=1.54-9.82] and good knowledge on colostrum feeding [AOR=3.53, 95%, CI=1.56-7.96] were found to be associated with practice of colostrum feeding. Conclusion: In the present study, mothers who had good practice of colostrum feeding is low. Socio-demographic, obstetrics and knowledge related factors were associated with practice of colostrum feeding. Promoting antenatal care visit and maternal education on breast feeding is recommended for the increasing of colostrum feeding practice of the mothers.