Identification of nucleated cells in urine using lectin staining

Microscopic examination of urinary sediment is an integral component in the evaluation of nephropathies, However, identification and differentiation o f the nucleated nonsquamous cells in urine is often difficult using such co nventional techniques as phase contrast or bright field microscopy, even af ter Papanicolaou staining, and requires a lot of experience. We now report a method to differentiate urinary cell types using lectin staining. Twenty- five lectins were examined with respect to their binding pattern on cryosec tions of the human kidney and urinary tract, as well as binding to blood ce lls. The specificity of lectin binding to a cell type both in situ and in u rine was confirmed by double labeling with specific antibodies directed aga inst various sections of the nephron or nucleated blood cells. For urine cy tologic examinations, acetone-fixed cytopreparations of urinary sediments w ere incubated with a combination of a fluorescein isothiocyanate (FITC)-cou pled and a rhodamine-coupled lectin, followed by staining of the nuclei wit h 4',6-diamidino-2-phenylindole. Specimens were examined in triple immunofl uorescence (FITC/rhodamine/UV). Cell types could be identified by their cha racteristic lectin-binding pattern. For example, the lectin combination of Sophora japonica agglutinin (aggl; SJA) and Erythrina cristagalli aggl (ECA ) permitted a differentiation between cells of the proximal tubules (SJA po sitive [SJA+], ECA+), distal tubules (SJA negative [SJA-], ECA+), collectin g ducts (SJA+, ECA-), and lymphocytes (SJA-, ECA-). In preliminary studies, examination Of urinary sediment in various chronic nephropathies by this t echnique Showed differences in their cellular excretion pattern. In summary , staining urinary sediments with combinations of lectins provides a rapid and relatively inexpensive method for a facilitated and reliable differenti ation of the various nucleated cell types in urine, (C) 2001 by the Nationa l Kidney Foundation, Inc.