Background: Acute kidney injury (AKI) is common in hospitalized patients and is associated with increased morbidity, mortality, and cost. Currently, AKI is diagnosed after symptoms manifest; available diagnostic tests (e.g., serum creatinine, urine microscopy, urine output) have limited ability to identify subclinical AKI. Because of the lack of treatment strategies, AKI typically is managed with supportive measures. However, strategies exist that may prevent renal insults in critically ill patients; therefore, early recognition of AKI is crucial for minimizing damage propagation. Content: Experimental and clinical studies have identified biomarkers that may facilitate earlier recognition of AKI or even identify patients at risk of AKI. Such biomarkers might aid in earlier implementation of preventive strategies to slow disease progression and potentially improve outcomes. This review describes some of the most promising novel biomarkers of AKI, including neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule 1 (KIM-1), interleukin 18 (lL-18), liver-type fatty-acid-binding protein (L-FABP), insulin-like-growth-factor-binding protein 7 (IGFBP7), and tissue inhibitor of metalloproteinase 2 (TIMP-2). Summary: We discuss biomarker test characteristics, their strengths and weaknesses, and future directions of their clinical implementation. IMPACT STATEMENT Acute kidney injury (AKI) affects millions of people in the US and increases morbidity, mortality, and healthcare expenditures. Available laboratory tests for early and accurate diagnosis of AKI have significant limitations. Novel biomarkers that predict development of AKI earlier are critically needed because they allow implementation of preventive strategies that potentially improve clinical outcomes. In this report, we review the most promising biomarkers of AKI that have been investigated during the past 2 decades and present a model for clinical implementation and areas for future investigation.