Fluorescent Lectin Labeling Protocol

The six fluorescein-labeled lectins Galanthus nivalis lectin (GNL; n = 8), Lens culinaris agglutinin (LCA; n = 5), Peanut agglutinin (PNA; n = 8), Solanum tuberosum lectin (STL; n = 8), Ulex europaeus agglutinin 1 (UEA I; n = 8), and Wheat germ agglutinin (WGA; n = 8) were purchased from Vector Laboratories (Burlingame, CA, USA). The lectin working solutions were prepared by diluting the stock solutions with simulated nasal electrolyte solution (SNES) to a final lectin concentration of 500 nmol/L. The lectin concentration was chosen based on a study by Engleder, Demmerer, Wang, Honeder, Zhu, Studenik, Wirth, Arnoldner, and Gabor [16 (link)] in a guinea pig model. The SNES contained 7.45 mg/mL of sodium chloride, 1.29 mg/mL of potassium chloride, and 0.32 mg/mL of calcium chloride dihydrate in distilled water [30 (link)]. All chemicals used for the preparation of SNES were of analytical grade and purchased from Carl Roth GmbH + Co. KG (Karlsruhe, Germany). This solution also served as a washing solution in the experiment. A solution of fluorescein isothiocyanate-conjugated bovine serum albumin (F-BSA; F/P ratio = 7–12; Sigma-Aldrich Corporation, St. Louis, MO, USA) in SNES (500 nmol/L) was used to rule out any non-specific protein binding. For lectin properties (source, molecular weight, binding motif, and molar fluorescein/protein (F/P) ratio), see Table 1.