Urine collected from diapers can be used for 2-D PAGE in infants and young children.
Urinary proteomic profiling has potential to identify candidate biomarkers of renal injury in infants provided an adequate urine sample can be obtained. Although diapers are used to obtain urine for clinical evaluation, their use for proteomic analysis has not been investigated. We therefore performed feasibility studies on the use of diaper-extracted urine for 2-D PAGE. Pediatric waste urine (2-20 mL) was applied to gel-containing, non-gel and cotton-gauze diapers and then mechanically expressed. Urine volume and total protein were measured pre- and post-extraction. Proteins were separated via 2-D PAGE following application of urine (20-40 mL) to each matrix. 2-D PAGE was also performed on clinical specimens collected using each diaper type. Differences in the adsorption and retention of urine volume and protein were noted between matrices. Non-gel and cotton-gauze diapers provided the best protein/volume recovery and the lowest interference with the Bradford assay. 2-D PAGE was also successfully completed using urine samples from both cotton fiber matrices. Conversely, samples from low-gel diapers demonstrated poor protein separation and reproducibility. Diapers containing cotton-fiber matrices appear adequate for 2-D PAGE. Qualitative and quantitative analyses of resolved proteins using replicate, high-resolution gels will be required, however, before diaper-extracted urine can be applied in proteomic profiling.
Department of Pediatrics, Kosair Charities Pediatric Clinical Research Unit, School of Medicine, University of Louisville, Louisville, KY, USA; Department of Pharmacology and Toxicology, School of Medicine, University of Louisville, Louisville, KY, USA. mjkennedy@vcu.edu.
Mary Jayne
MJ
Kennedy
Angela
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Griffin
Ruifeng
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Su
Michael
M
Merchant
Jon
J
Klein