title: Staining histological lung sections with Sudan Black B or Sudan III for automated identification of alveolar epithelial type II cells.
abstract: Alveolar epithelial type II (AE2) cells produce, store and secrete pulmonary surfactant and serve as progenitor cells for the alveolar epithelium. They are thus an interesting target in wide fields of pulmonary research. Stereological methods allow their quantification based on measurements on histological sections. A proper AE2 cell quantification, however, requires a method of tissue processing that results in little tissue shrinkage during processing. It was recently shown that a primary fixation with a mixture of glutaraldehyde and formaldehyde, postfixation with osmium tetroxide and uranyl acetate and embedding in glycol methacrylate fulfills this requirement. However, a proper quantification, furthermore, requires a secure identification of the cells under the microscope. Classical approaches using routine stainings, high magnifications and systematic uniform random sampling can result in a tedious counting procedure. In this article we show that Sudan Black B and Sudan III staining in combination with the previously described "low shrinkage method" of tissue processing result in good staining of lamellar bodies of AE2 cells (their storing organelles of surfactant) and thus provide a good signal of AE2 cells, which allows their easy and secure identification even at rather low magnifications. We further show that this signal enables automated detection of AE2 cells by image analysis, which should make this method a suitable staining method for the recently developed and more efficient proportionator sampling.
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Carl-Neuberg-Straße 1, Hannover 30625, Germany; Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany; REBIRTH Cluster of Excellence, Hannover, Germany. Electronic address: schneider.jan@mh-hannover.de.
authors.forename: Jan Philipp
authors.initials: JP
authors.lastname: Schneider
authors.affiliation: Visiopharm, Agern Allé 3, Hoersholm, Denmark. Electronic address: lap@visiopharm.com.
authors.forename: Lars
authors.initials: L
authors.lastname: Pedersen
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Carl-Neuberg-Straße 1, Hannover 30625, Germany; Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany; REBIRTH Cluster of Excellence, Hannover, Germany. Electronic address: muehlfeld.christian@mh-hannover.de.
authors.forename: Christian
authors.initials: C
authors.lastname: Mühlfeld
authors.affiliation: Institute of Functional and Applied Anatomy, Hannover Medical School, Carl-Neuberg-Straße 1, Hannover 30625, Germany; Biomedical Research in Endstage and Obstructive Lung Disease Hannover (BREATH), Member of the German Center for Lung Research (DZL), Hannover, Germany; REBIRTH Cluster of Excellence, Hannover, Germany. Electronic address: ochs.matthias@mh-hannover.de.
authors.forename: Matthias
authors.initials: M
authors.lastname: Ochs
mesh_terms: D056809:Alveolar Epithelial Cells / Q000473:pathology; D000818:Animals; D001391:Azo Compounds / Q000737:chemistry; D001499:Bayes Theorem; D004396:Coloring Agents / Q000737:chemistry; D006801:Humans; D007090:Image Interpretation, Computer-Assisted; D008168:Lung / Q000473:pathology*; D008171:Lung Diseases / Q000473:pathology; D051379:Mice; D009281:Naphthalenes / Q000737:chemistry; D013194:Staining and Labeling
chemical_list: D001391:Azo Compounds; D004396:Coloring Agents; D009281:Naphthalenes; C016118:Sudan Black B; C033006:sudan III
keywords: Alveolar epithelial type II cells; Lamellar bodies; Lung; Stereology; Sudan Black B; Sudan III