Background: New urinary biomarkers like neutrophil gelatinase-associated lipocalin (NGAL), liver-type fatty acid binding protein (L-FABP), and kidney injury molecule-1 (KIM-1) open the opportunity to detect kidney injuries in early stages. Our study aimed at evaluating NGAL, L-FABP, and KIM-1 in comparison to established markers of urine protein differentiation for detection of renal dysfunction. Methods: On the basis of the PROTIS expert system (for differentiation of glomerulo-/tubulopathy) urine and plasma samples of 263 randomly selected patients were routinely examined (urine: total protein, albumin, IgG, α 1 -microglobulin, creatinine, and dip stick results for leukocytes, blood, protein, glucose, pH, and nitrite; plasma: creatinine and cystatin C) followed by the analysis of the new urine biomarkers NGAL (CMIA), L-FABP (ECLIA), and KIM-1 (ELISA). Results: Of the three new markers L-FABP showed the highest correlation with α 1 -microglobulin (r=0.76, p<0.01) and was closest associated with the degree of tubular proteinuria assessed by the PROTIS system. NGAL distinguished the PROTIS proteinuria groups with distinctive tubular proteinurias from the controls as well, but revealed a marked diagnostic influence by leukocyturia. Urinary KIM-1 revealed only a weak diagnostic value for the detection of renal injury. Conclusions: Urinary NGAL and L-FABP proved to be promising candidates for detecting injuries of the renal tubular system over a broad range of clinical conditions. L-FABP showed a better diagnostic performance and a lower interference by leukocyturia and hematuria than NGAL. Both markers may serve as sensitive tissue injury markers in addition to the established markers of renal functional impairment.