Background: Most validations of paediatric glomerular filtration rate (GFR) estimating equations using standardized creatinine (CR) and cystatin C (CYS) assays have comprised relatively small cohorts, which makes accuracy across subgroups of GFR, age, body mass index (BMI) and gender uncertain. To overcome this, a large cohort of children referred for GFR determination has been established from several European medical centres. Methods: Three thousand four hundred eight measurements of GFR (mGFR) using plasma clearance of exogenous substances were performed in 2218 children aged 2–17 years. Validated equations included Schwartz-2009 CR /2012 CR/CYS/CR+CYS , FAS CR/CYS/CR+CYS , LMR CR , Schwartz-Lyon CR , Berg CYS , CAPA CYS , CKD-EPI CYS , Andersen CR+CYS and arithmetic means of the best single-marker equations in explorative analysis. Five metrics were used to compare the performance of the GFR equations: bias, precision and three accuracy measures including the percentage of GFR estimates (eGFR) within ± 10% (P 10 ) and ± 30% (P 30 ) of mGFR. Results: Three of the cystatin C equations, Berg CYS , CAPA CYS and CKD-EPI CYS , exhibited low bias and generally satisfactory accuracy across all levels of mGFR; CKD-EPI CYS had more stable performance across gender than the two other equations. Among creatinine equations, Schwartz-Lyon CR had the best performance but was inaccurate at mGFR < 30 mL/min/1.73 m 2 and in underweight patients. Arithmetic means of the best creatinine and cystatin C equations above improved bias compared to the existing composite creatinine+cystatin C equations. Conclusions: The present study strongly suggests that cystatin C should be the primary biomarker of choice when estimating GFR in children with decreased GFR. Arithmetic means of well-performing single-marker equations improve accuracy further at most mGFR levels and have practical advantages compared to composite equations.