• Heparin-binding protein could be a promising biomarker that distinguish severe sepsis from sepsis in the pediatric intensive care unit. • The combination of heparin-binding protein with procalcitonin and lactate could significantly improve the diagnostic value for severe sepsis. • Heparin-binding protein levels are related to the severity of the disease among children with sepsis. • The dynamic changes of heparin-binding protein at 72 h after enrollment could predict in-hospital mortality among children with sepsis in the pediatric intensive care unit. The aim of this study is to assess Heparin-binding protein (HBP) as a diagnostic and prognostic biomarker of severe sepsis in the pediatric intensive care unit (PICU). A multicenter, prospective study was conducted among children with sepsis in nine PICUs in China from October 2019 to June 2021. Plasma levels of HBP, procalcitonin (PCT), C-reactive protein (CRP), lactate, and white blood cell (WBC) count were determined at enrollment and 72 h after enrollment. Of 355 included patients, 132 patients were diagnosed with non-severe sepsis (referred to as sepsis), 223 patients had severe sepsis. Patients with severe sepsis had significantly elevated levels of HBP compared with sepsis (median 170.5 vs. 74.1 ng/mL, P < 0.001). Adding HBP to a diagnostic model with PCT and lactate could significantly improve the diagnostic capability for severe sepsis. The plasma levels of HBP correlated positively with the number of dysfunctional organs. After adjusting for confounding factors, the declined levels of HBP at 72 h had a significant association with decreased in-hospital mortality (adjusted odds ratio (aOR) 0.242, P < 0.001). The levels of HBP showed weak positive correlations with PCT, CRP, WBC, and no correlation to lactate. HBP at enrollment can be an independent indicator for severe sepsis and the dynamic changes at 72 h can be a predictor for in-hospital mortality in PICU. [ABSTRACT FROM AUTHOR]