Background: Cryptosporidiosis is an intestinal infection caused by an intracellular protozoan parasite Cryptosporidium; which is one of the critical causes of diarrhea in children around the world. Objective: to detect Cryptosporidium infection in clinical samples from pediatrics suffering from diarrhea in Misurata, Libya, and evaluate three methods of diagnosis. Methods: 150 stool samples were collected from children with diarrhea, in the age group (1 month – 15 years) in both genders who attended Misurata Central Hospital. The stool samples were inspected by microscopic examination Modified Ziehl Neelsen (MZN), crypto-strip test and enzyme-linked immunosorbent assay (ELISA). Results: indicated that 17 children in the study group (11.3%) were positive for Cryptosporidium infection using MZN, while 19 (12.7%) were positive using the Crypto-strip test and ELISA, bearing in mind that there was no significant difference between male and females nor between urban and rural areas. The higher infection appeared in age group < 5-year-old, the sensitivity, specificity of the Crypto-strip test and ELISA were 89.47%, 100% respectively. Conclusions: The high prevalence rate of Cryptosporidium infection among children of less than 5 years old in both genders were sensitive to be infected with Cryptosporidium in both rural and urban areas, the three methods proved to be a useful mean for diagnosis of cryptosporidiosis in stool samples.