In Europe from 2006 and nowadays all over the world, there are restrictions about the addition of antibiotics to calves' food in order to minimize the rates of bacterial resistance to antibiotics. Thus, this study was conducted to determine the anti-diarrheal effect of probiotic supplementation in neonatal buffalo calves at a dose of 10 g for each animal once daily for three weeks comparing with non-supplemented control group that fed only on milk replacer. Fecal score, selective hematological and biochemical studies were analyzed to evaluate the effect of probiotics on the frequency of diarrhea caused by Escherichia coli (E. coli) and the Probability of having any side effects on blood picture  and liver function. Results revealed that the probiotics mixture used at this study had no significant changes at the values of red and white blood cells, liver enzymes, glucose and electrolytes. however, a  significant decrease in the count of fecal  E. coli wihout diarrhea was observed all over the experimental periods compared to the non-supplemented animals. It could be concluded that the addition of probiotics to calves food at the period before weaning is safe and achieving antidiarreal and antimicrobial benefits.