Description of an Automated Method for Urea Nitrogen Determination in Bronchoalveolar Lavage Fluid (BALF) of Neonates and Infants

Abstract

Bronchoalveolar lavage (BAL) partially recovers both the instilled saline and the alveolar fluid, so-called epithelial lining fluid (ELF), but a correction for the dilution due to the BAL technique itself is needed to know the amount of recovered ELF. In this regard, urea nitrogen may be useful and has been proposed to calculate ELF. The aim of the present study was to develop and validate a new method to measure urea nitrogen in BAL fluid (BALF). We used 19 BALF samples obtained from neonates and infants with different respiratory conditions. The urea nitrogen assay was carried out on Cobas c311 analyzer (Roche Diagnostics). A validation study shows that the method is perfectly linear (R(2) = 0.999), sensitive (limit of detection = 0.055 mg/dL; limit of quantification = 0.16 mg/dL), repeatable (low = 0.15 ± 0.02, 13.3%; high = 1.80 ± 0.02, 1.1%), reproducible (low = 0.14 ± 0.02, 14.2 %; high = 1.76 ± 0.04, 2.2 %) with accuracy ranging between 93-96%. Our results support the robustness of validated procedure since the described method appears simple, precise, rapid, and suitable for routine analysis. Thus, it may be used to correct concentration of various noncellular BAL components and calculate their ELF amounts in neonates and infants.

Clinical utility of bronchoalveolar lavage cytomegalovirus viral loads in the diagnosis of cytomegalovirus pneumonitis in infants

“…A possible reason for this was that a manual plate‐based method of urea determination was used, introducing a further variability in the final CMV quantification in ELF. An automated method should be utilized in future studies of ELF [Pocino et al, ].…”

Heparin-binding protein, lysozyme, and inflammatory cytokines in bronchoalveolar lavage fluid as diagnostic tools for pulmonary infection in lung transplanted patients

“…To adjust for dilution of BALF in estimating the biomarker concentrations in the lung epithelial lining fluid, we analyzed urea in BALF and concomitant plasma samples with a QuantiChrom Urea Assay Kit (BioAssay Systems, Hayward, CA) according to the protocol provided by the manufacturer. The ratio [urea in plasma]/[urea in BALF] was used as a coefficient for dilution to adjust biomarker levels as described previously by Pocino et al …”

Preventing lung pathology and mortality in rabbit Staphylococcus aureus pneumonia models with cytotoxin-neutralizing monoclonal IgGs penetrating the epithelial lining fluid

“…In parallel, anti-human and anti-rabbit IgG reagents were also used in ELISA to measure total human and rabbit IgG levels. Urea concentrations of BALF and serum samples were determined to normalize for the dilution of BALF samples and to calculate the real IgG concentrations in the epithelial lining fluid (ELF) of the single matched time point samples 29–31 .…”