The aim of the study was to create a specific pattern of protein expression changes in the plasma of 2-week-old calves treated with excessive lactose doses. Identification of proteins, the expression of which changes during the short-term administration of lactose, may help to exclude or confirm the causes of diarrhea observed in newborn calves. The present study utilized 2-D electrophoresis combined with MALDI-TOF mass spectrometry and identified 6 blood plasma proteins, the expression of which changed under the influence of lactose added to the milk replacer. The pattern of expression of blood plasma proteins, including decreased expression of fibrinogen and apolipoprotein A-IV and increased expression of alpha-1B-glycoprotein, alpha-1-antiproteinase, and apolipoprotein E in 2-week-old calves fed with lactose-rich milk replacer, can be used to determine the causes of diarrhea in calves.