118 - Urine collection in cotton wool balls yields similar concentrations of albumin, creatinine, uromodulin, exosomal proteins, compared to clean catch samples

Poster Number: 118
Publication Number: 118.125

Eveline Staub, Royal North Shore Hospital, University of Sydney, St Leonards, New South Wales, Australia; Qinghua Cao, Kolling Institute of Medical Research, Sydney, New South Wales, Australia; Xin-Ming Chen, The University of Sydney, Sydney, New South Wales, Australia; Carol Pollock, Royal North Shore Hospital, University of Sydney, Sydney, New South Wales, Australia

Background: Collecting urine samples in neonates by catheterization or suprapubic puncture cause trauma, while self-adhesive collection bags can damage fragile skin. Collecting samples from urine-soaked cotton wool balls placed in diapers is an alternative, but could yield false analysis results when substances are retained within the cotton wool fibres.

Objective: To compare the concentration of albumin, creatinine, neutrophil gelatinase-associated lipocalin (NGAL), uromodulin and exosomal protein between clean catch urine samples and samples collected in cotton wool balls in preterm and term neonates.

Design/Methods: Clean catch urine samples were collected from 19 patients in the local neonatal intensive care unit and divided into three portions: standard sample (clean catch urine), sample 2 (cotton wool balls soaked with a portion of standard sample, then urine immediately expressed), and sample 3 (cotton wool balls soaked with a portion of standard sample, placed in diaper and kept in an incubator at 37˚C for 2 hours, then urine expressed). The samples were then analysed for albumin, creatinine, NGAL, uromodulin, bicinchoninic acid (BCA, surrogate marker for levels of exosomal protein) by commercially available ELISA kits. Differences in substance concentration between standard sample and sample 2 or sample 3 were expressed as absolute differences and percentage of median concentration. Exosomes were extracted from all three samples of one patient for visualization under electron microscopy.

Results: Urine samples of 19 infants (10 males, median gestational age 34+3 wks, range 25+0 - 41+1) were collected at a median age of 19 days (range 2-95). Concentration differences between standard sample and sample 2 were: albumin -0.12µg/ml (-1.9%), creatinine -1.7mg/dl (-7.3%), NGAL 0ng/ml (0%), uromodulin -0.09µg/ml (-1.6%), BCA -0.03mg/ml (-4.5%). Differences between standard sample and sample 3 were: albumin +0.002µg/ml (+0.02%), creatinine -1.7mg/dl (-7.5%), NGAL -2.4ng/ml (-33%), uromodulin 0µg/ml (0%), BCA 0.04mg/ml (+7.3%). Exosomes were visualized under electron microscopy in all three types of urine samples (Fig 1).  Conclusion(s): Neonatal urine samples collected in cotton wool balls yield similar concentrations of albumin, creatinine, uromodulin, even when incubated for up to 2 hours in typical temperatures of incubators, with differences in concentrations generally less than 10%. Larger urine components such as exosomes do not get lost in the fibres of cotton wool balls, as shown by similar exosomal protein content and visualization of exosomes under electron microscopy in all three sample types.
Figure 1. Urine exosomes from three urine sample types visualized by electron microscopyExosomes were extracted from three different urine sample types of preterm and term neonates: A: clean catch urine; B: urine collected in cotton wool balls and immediately expressed; C: urine collected in cotton wool balls, placed within a diaper in an incubator heated to 37∘C for 2 hours. D: positive control from cell culture supernatant. Exosomes measuring 39 to 95nm in diameter, black reference bars are 200nm in A and D, 100nm in B and C.