Strategy and rationale for urine collection protocols employed in the NEPTUNE study

Area of Science:

• Nephrology
• Biomarker Discovery
• Translational Research

Background:

• Glomerular diseases pose significant health risks, necessitating effective monitoring and intervention.
• Urine analysis offers a non-invasive approach to identify kidney-specific molecular signatures.
• Development of non-invasive biomarkers is crucial for managing glomerular diseases.

Purpose of the Study:

• To describe the urine procurement and banking procedures within the Nephrotic Syndrome Study Network (NEPTUNE).
• To establish standardized methods for collecting urine for biomarker discovery in nephrotic syndrome.
• To support the identification of non-invasive biomarkers for minimal change disease, focal segmental glomerulosclerosis, and membranous nephropathy.

Main Methods:

• Standardized urine collection across multiple North American centers.
• Detailed protocols for urine procurement and banking, including storage conditions and preservative use.
• Evaluation of analyte stability (NGAL, retinol binding globulin, alpha-1 microglobulin) under various conditions.

Main Results:

• The NEPTUNE protocol enables robust detection of proteins and RNAs from urine.
• Analyte performance was assessed with and without preservatives (thymol, toluene, boric acid).
• High-quality RNA and protein were successfully recovered from urine cellular pellets and exosomes.

Conclusions:

• The NEPTUNE urine collection protocol supports longitudinal monitoring over 5 years.
• This standardized approach yields a valuable resource for validating glomerular disease biomarkers.
• The protocol facilitates the exploration of novel and established markers using comprehensive clinical and histopathologic data.