title: Visualization of Nanofibrillar Cellulose in Biological Tissues Using a Biotinylated Carbohydrate Binding Module of β-1,4-Glycanase.
abstract: Nanofibrillar cellulose is a very promising innovation with diverse potential applications including high quality paper, coatings, and drug delivery carriers. The production of nanofibrillar cellulose on an industrial scale may lead to increased exposure to nanofibrillar cellulose both in the working environment and the general environment. Assessment of the potential health effects following exposure to nanofibrillar cellulose is therefore required. However, as nanofibrillar cellulose primarily consists of glucose moieties, detection of nanofibrillar cellulose in biological tissues is difficult. We have developed a simple and robust method for specific and sensitive detection of cellulose fibers, including nanofibrillar cellulose, in biological tissue, using a biotinylated carbohydrate binding module (CBM) of β-1,4-glycanase (EXG:CBM) from the bacterium Cellulomonas fimi. EXG:CBM was expressed in Eschericia coli, purified, and biotinylated. EXG:CBM was shown to bind quantitatively to five different cellulose fibers including four different nanofibrillar celluloses. Biotinylated EXG:CBM was used to visualize cellulose fibers by either fluorescence- or horse radish peroxidase (HRP)-tagged avidin labeling. The HRP-EXG:CBM complex was used to visualize cellulose fibers in both cryopreserved and paraffin embedded lung tissue from mice dosed by pharyngeal aspiration with 10-200 μg/mouse. Detection was shown to be highly specific, and the assay appeared very robust. The present method represents a novel concept for the design of simple, robust, and highly specific detection methods for the detection of nanomaterials, which are otherwise difficult to visualize.
authors.affiliation: †National Research Centre for the Working Environment, Lersø Parkallé 105, DK-2100 Copenhagen Ø, Denmark.
authors.forename: Kristina Bram
authors.initials: KB
authors.lastname: Knudsen
authors.affiliation: ‡Section for Biomolecular Sciences, Department of Biology, University of Copenhagen, Copenhagen, DK-2200 N, Denmark.
authors.forename: Christian
authors.initials: C
authors.lastname: Kofoed
authors.affiliation: ‡Section for Biomolecular Sciences, Department of Biology, University of Copenhagen, Copenhagen, DK-2200 N, Denmark.
authors.forename: Roall
authors.initials: R
authors.lastname: Espersen
authors.affiliation: ‡Section for Biomolecular Sciences, Department of Biology, University of Copenhagen, Copenhagen, DK-2200 N, Denmark.
authors.forename: Casper
authors.initials: C
authors.lastname: Højgaard
authors.affiliation: ‡Section for Biomolecular Sciences, Department of Biology, University of Copenhagen, Copenhagen, DK-2200 N, Denmark.
authors.forename: Jakob Rahr
authors.initials: JR
authors.lastname: Winther
authors.affiliation: ‡Section for Biomolecular Sciences, Department of Biology, University of Copenhagen, Copenhagen, DK-2200 N, Denmark.
authors.forename: Martin
authors.initials: M
authors.lastname: Willemoës
authors.affiliation: §Stora Enso, FI-00100 Helsinki, Finland.
authors.forename: Irene
authors.initials: I
authors.lastname: Wedin
authors.affiliation: ∥UPM Kymmene Oyj, FI-00100 Helsinki, Finland.
authors.forename: Markus
authors.initials: M
authors.lastname: Nuopponen
authors.affiliation: ⊥Nanosafety Research Centre, Finnish Institute of Occupational Health, P.O. Box 40, FI-00250 Helsinki, Finland.
authors.forename: Sara
authors.initials: S
authors.lastname: Vilske
authors.affiliation: ⊥Nanosafety Research Centre, Finnish Institute of Occupational Health, P.O. Box 40, FI-00250 Helsinki, Finland.
authors.forename: Kukka
authors.initials: K
authors.lastname: Aimonen
authors.affiliation: †National Research Centre for the Working Environment, Lersø Parkallé 105, DK-2100 Copenhagen Ø, Denmark.
authors.forename: Ingrid Elise Konow
authors.initials: IE
authors.lastname: Weydahl
authors.affiliation: ⊥Nanosafety Research Centre, Finnish Institute of Occupational Health, P.O. Box 40, FI-00250 Helsinki, Finland.
authors.forename: Harri
authors.initials: H
authors.lastname: Alenius
authors.affiliation: ⊥Nanosafety Research Centre, Finnish Institute of Occupational Health, P.O. Box 40, FI-00250 Helsinki, Finland.
authors.forename: Hannu
authors.initials: H
authors.lastname: Norppa
authors.affiliation: ⊥Nanosafety Research Centre, Finnish Institute of Occupational Health, P.O. Box 40, FI-00250 Helsinki, Finland.
authors.forename: Henrik
authors.initials: H
authors.lastname: Wolff
authors.affiliation: †National Research Centre for the Working Environment, Lersø Parkallé 105, DK-2100 Copenhagen Ø, Denmark.|#Institute of Public Health, Copenhagen University, Øster Farimagsgade 5, Copenhagen K, Denmark.
authors.forename: Håkan
authors.initials: H
authors.lastname: Wallin
authors.affiliation: †National Research Centre for the Working Environment, Lersø Parkallé 105, DK-2100 Copenhagen Ø, Denmark.|○Department of Micro- and Nanotechnology, DTU, Kgs. Lyngby, Denmark.
authors.forename: Ulla
authors.initials: U
authors.lastname: Vogel
mesh_terms: D000818:Animals; D019921:Biotinylation; D002482:Cellulose / Q000378:metabolism* / Q000648:ultrastructure*; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D007150:Immunohistochemistry; D008168:Lung / Q000166:cytology; D051379:Mice; D057139:Nanofibers / Q000648:ultrastructure*; D011485:Protein Binding; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D002482:Cellulose; D006026:Glycoside Hydrolases; C020668:glycanase