Aflatoxin M1 (AFM1) concentrations were determined in raw and UHT cow milk samples collected in different regions of Bosnia and Herzegovina and Croatia during the autumn months of 2014. The mean AFM1 levels in the raw milk samples were (ng/kg): 6.22 in Bosnia and Herzegovina, 5.65 in Croatia. In all except one milk sample, AMF1 levels were below the LOQ value of 34.2 ng/kg (ELISA method). In four milk samples, AFM1 concentrations exceeded the EU MRL of 50 ng/kg. Samples were subjected to LC-MS/MS analysis which confirmed elevated values determined by ELISA. Elevated levels were in the range 56.6–132.6 ng/kg. Two positive milk samples from Bosnia and Herzegovina originated from Una Sana Canton, two from Croatia from eastern Croatia. The highest AFM1 levels of 132.6 ng/kg was measured in milk from eastern Croatia. In 214 samples of processed UHT milk from Bosnia and Herzegovina and Croatia, AFM1 ranged from 2.29 ng/kg to 21.4 ng/kg, all below the LOQ value. AFM1 exceeded the EU MRL value in only 0.62% of milk samples, indicating the sporadic use of contaminated feedstuff at farms in both countries. •Raw and UHT milk were collected in Bosnia and Herzegovina and Croatia.•During autumn 2014 only 0.62% raw milk exceeded the maximum EU limit for AFM1.•Two samples from Bosnia and Herzegovina and two from Croatia exceeded EU limit.•Four elevated AFM1 concentrations were confirmed by LC-MS/MS.•In all UHT milk samples AFM1 levels were below LOQ of the ELISA method.