Early diagnosis of acute kidney injury by aptasensors

DESCRIPTION provided by applicant Acute kidney injury AKI affects more than million people worldwide each year with reported mortalities from to in different patient populations However methods are lacking to support the early diagnosis of AKI that could lead to early intervention improved therapy better prognosis and lower medical costs For these reasons the development of early diagnosis tools for AKI is one of the solicited research topics listed by the NIDDK Neutrophil gelatinase associated lipocalin NGAL is a reliable biomarker for AKI and may also have some predictive uses We propose to develop a portable device for reliable early detection of AKI at point of care The innovation of our proposal lies in the combination of aptamer technology with microcantilever detection of NGAL an early marker of AKI h at the onset of AKI cutoff ng ml Aptamers are single stranded DNAs or RNAs that bind their targets avidly and specifically and are selected in vitro by Systematic Evolution f Ligands by Exponential enrichment SELEX For biosensor development aptamers have many advantages over antibodies including their smaller size amenability to chemical synthesis and modifications and adaptability to a broad range of assay formats including a microcantilever detection system A microcantilever is a beam often of silicon that is m thick and m long and anchored only at one end The microcantilever bends when its surface stress changes due to the conformation change that occurs upon ligand receptor e g aptamer target interaction on its surface The differential surface stress between a reference cantilever coated with a scrambled nucleic acid that does not bind the analyte and a sensing cantilever coated with an aptamer that binds the analyte is measured by interferometry In a phase I study we isolated several aptamers that bind NGAL and demonstrated the abilities of one of these to sense NGAL when the aptamer is attached to a microcantilever In this phase II study we propose the following specific aims Compare several aptamers for specificity functionality in the presence of serum proteins affinity for NGAL effectiveness in the presence of salt mixtures characteristic of plasma and urine and compatibility with the microcantilever device Optimize the best performing aptamers for length sequence stability and optimal assay performance in blood and urine and Optimize the microcantilever device for aptamer sensing dynamic range of nM for detection signal noise ratio andgt covariance andlt Miniaturize the microcantilever device to a size equal to or smaller than a shoebox From these studies we expect to develop an optimized and reliable aptamer functionalized microcantilever by which NGAL can be quantified in plasma and urine samples with robust signal noise ratio sensitivity detection limit and dynamic range of measurement Phase III studies will involve manufacturing and validating the device for monitoring NGAL to detect AKI at point of care PUBLIC HEALTH RELEVANCE Acute kidney injury AKI affects more than million people worldwide per year with mortality However methods for early diagnosis of AKI are still lacking and consequently this is one of the research topics solicited by the National Institute of Diabetes and Digestive and Kidney Diseases We propose to develop aptamer based biosensors for early diagnosis of AKI at point of care based on NGAL an early marker as early as hours for AKI