title: Dissociation of skeletal muscle for flow cytometric characterization of immune cells in macaques.
abstract: The majority of vaccines and several treatments are administered by intramuscular injection. The aim is to engage and activate immune cells, although they are rare in normal skeletal muscle. The phenotype and function of resident as well as infiltrating immune cells in the muscle after injection are largely unknown. While methods for obtaining and characterizing murine muscle cell suspensions have been reported, protocols for nonhuman primates (NHPs) have not been well defined. NHPs comprise important in vivo models for studies of immune cell function due to their high degree of resemblance with humans. In this study, we developed and systematically compared methods to collect vaccine-injected muscle tissue to be processed into single cell suspensions for flow cytometric characterization of immune cells. We found that muscle tissue processed by mechanical disruption alone resulted in significantly lower immune cell yields compared to enzymatic digestion using Liberase. Dendritic cell subsets, monocytes, macrophages, neutrophils, B cells, T cells and NK cells were readily detected in the muscle by the classic human markers. The methods for obtaining skeletal muscle cell suspension established here offer opportunities to increase the understanding of immune responses in the muscle, and provide a basis for defining immediate post-injection vaccine responses in primates.
authors.affiliation: Department of Medicine, Karolinska Institutet, Stockholm, Sweden.|Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Frank
authors.initials: F
authors.lastname: Liang
authors.affiliation: Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Aurélie
authors.initials: A
authors.lastname: Ploquin
authors.affiliation: Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: José DelaO
authors.initials: JD
authors.lastname: Hernández
authors.affiliation: Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Hugues
authors.initials: H
authors.lastname: Fausther-Bovendo
authors.affiliation: Department of Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Gustaf
authors.initials: G
authors.lastname: Lindgren
authors.affiliation: Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Daphne
authors.initials: D
authors.lastname: Stanley
authors.affiliation: Laboratory of Pharmaceutics, University of the Basque Country, School of Pharmacy, Vitoria, Spain.
authors.forename: Aiala Salvador
authors.initials: AS
authors.lastname: Martinez
authors.affiliation: Laboratory of Molecular Microbiology, NIAID/NIH, Bethesda, MD.
authors.forename: Jason M
authors.initials: JM
authors.lastname: Brenchley
authors.affiliation: Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Koup
authors.affiliation: Department of Medicine, Karolinska Institutet, Stockholm, Sweden.|Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Karin
authors.initials: K
authors.lastname: Loré
authors.affiliation: Vaccine Research Center, NIAID/NIH, Bethesda, MD.
authors.forename: Nancy J
authors.initials: NJ
authors.lastname: Sullivan
mesh_terms: D000818:Animals; D003713:Dendritic Cells / Q000276:immunology; D005434:Flow Cytometry / Q000379:methods; D008214:Lymphocytes / Q000276:immunology; D008251:Macaca / Q000276:immunology*; D008264:Macrophages / Q000276:immunology; D009000:Monocytes / Q000276:immunology; D018482:Muscle, Skeletal / Q000276:immunology*; D009504:Neutrophils / Q000276:immunology; D014612:Vaccines / Q000276:immunology
chemical_list: D014612:Vaccines
keywords: Flow cytometry; Nonhuman primate; Skeletal muscle; Vaccine administration