Infection often occurs in children with malignant hematology and causes morbidity and mortality. Antibiotics should be given based on the results of culture and antibiotic susceptibility test. Ampicillin Sulbactam is a first-line drug in children with infection. The purpose of this study was to obtain an objective overview of Ampicillin Sulbactam resistance patterns of culture in patients hospitalized in pediatric Hemato-Oncology Ward, Dr Soetomo Hospital, Surabaya. This was a descriptive study. Data on culture and susceptibility test result to Ampicillin Sulbactam were taken from medical records of those patients between September 2012 - February 2013. There were 342 culture examinations in 88 patients (44 girls and 44 boys). Most of the children aged <5 years (58%). Growth of bacterial cultures was obtained in 83 namely Escherichia coli, Burkholderia cepacia, and Klebsiella oxytoca (Gram-negative) and coagulation negative Staphylococcus (CONS), Staphylococcus aureus, and Staphylococcus saprophyticus (Gram-positive) were found in blood culture. Staphylococcus aureus (Gram positive) and E. coli, Klebsiella pneumoniae, and B.cepacia (Gram-negatif) were found in urine. The result of antibiotic susceptibility test culture showed resistance to Ampicillin Sulbactam in E. coli (69%), Burkholderia cepacia (85%), CONS (50%), Staphylococcus aureus (25%), K. pneumoniae (85%), Klebsiella oxytoca (50%), and P. aeroginosa (67%). There was ampicillin sulbactam resistance in children hospitalized in pediatric Hemato-Oncology Ward, Dr Soetomo Hospital, Surabaya against E. coli (69%), Burkholderia cepacia (85%), CONS (50%), Staphylococcus aureus (25%), K.pneumoniae (85%), and Klebsiella oxytoca (50%), P. aeroginosa (67%).