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Hyperbilirubinemia is an important problem in the neonatal period. It is a cause of concern for nursesand physician and a source of anxiety for the parents. High bilirubin level may be toxic to the developing central nervous system and may cause neurological impairment even in term neonates. The aim of this study is to identify the factors affecting accuracy of measuring TcB level in the neonates with hyperbilirubinemia. A descriptive exploratory design was used in the study. Data were collected using the neonatal medical records and TcB measurement were done for two successive times for every neonate in subsequent two days during the routine times of measuring TSB according to the unit's policy. A purposeful sample of one hundred neonateswere included. The results of this study revealed that there was statistically significant difference between TcB and TSB. No statistically significant difference between TcB and TSB measurements before and after starting phototherapy, but it differed significantly during phototherapy. The current study concluded that measuring transcutaneous bilirubin level on admission, before and after phototherapy help in determining the serum bilirubin level.