title: Phenotypic characterization and functional analysis of human tumor immune infiltration after mechanical and enzymatic disaggregation.
abstract: Multi-parametric flow cytometry analysis is a reliable method for phenotypic and functional characterization of tumor infiltrating immune cells (TIIC). The isolation of infiltrating leukocytes from solid tumors can be achieved through various methods which can be both enzymatic and mechanical; however, these methods may alter cell biology. The aim of this study was to compare the effects of three tissue disaggregation techniques on TIIC biology in breast, kidney and lung tumor specimens. We therefore compared two enzymatic treatments using either collagenase type IA alone or in combination with collagenase type IV and DNase I type II, and one mechanical system (Medimachine™). We evaluated the impact of treatments on cell viability, surface marker integrity and proliferative capacity. We show that cell viability was not significantly altered by treatments. However, enzymatic treatments decreased cell proliferation; specifically collagenases and DNase provoked a significant decrease in detection of surface markers such as CD4, CD8, CD45RA and CD14, indicating that results of phenotypic studies employing these techniques could be affected. In conclusion, mechanical tissue disaggregation by Medimachine™ appears to be optimal to maintain phenotypic and functional TIIC features.
authors.affiliation: Research Center, Centre Hospitalier de l'Université de Montréal (CRCHUM), Hôpital Notre-Dame, Université de Montréal and Institut du Cancer de Montréal, Montréal, Québec, Canada.
authors.forename: Cécile
authors.initials: C
authors.lastname: Grange
authors.forename: Jason
authors.initials: J
authors.lastname: Létourneau
authors.forename: Marie-Andrée
authors.initials: MA
authors.lastname: Forget
authors.forename: Jessica
authors.initials: J
authors.lastname: Godin-Ethier
authors.forename: Jocelyne
authors.initials: J
authors.lastname: Martin
authors.forename: Moishe
authors.initials: M
authors.lastname: Liberman
authors.forename: Mathieu
authors.initials: M
authors.lastname: Latour
authors.forename: Hugues
authors.initials: H
authors.lastname: Widmer
authors.forename: Jean-Baptiste
authors.initials: JB
authors.lastname: Lattouf
authors.forename: Ciriaco A
authors.initials: CA
authors.lastname: Piccirillo
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Cailhier
authors.forename: Réjean
authors.initials: R
authors.lastname: Lapointe
mesh_terms: D001943:Breast Neoplasms / Q000276:immunology*; D049109:Cell Proliferation; D002469:Cell Separation / Q000379:methods*; D005260:Female; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007680:Kidney Neoplasms / Q000276:immunology*; D008175:Lung Neoplasms / Q000276:immunology*; D016246:Lymphocytes, Tumor-Infiltrating / Q000166:cytology / Q000276:immunology*