Full Spectrum Flow Cytometry for High-Dimensional Immunophenotyping of Mouse Innate Lymphoid Cells

Abstract

Purpose and appropriate sample typesThis 25-parameter, 22-colour full spectrum flow cytometry panel was designed and optimised for the comprehensive enumeration and functional characterisation of innate lymphoid cell (ILC) subsets in mouse tissues (Table 1). The panel presented here allows the discrimination of ILC progenitors (ILCP), ILC1, ILC2, NCR+ ILC3, NCR- ILC3, CCR6+ lymphoid tissue-inducer (LTi)-like ILC3 and mature natural killer (NK) cell populations. Further characterisation of ILC and NK cell functional profiles in response to stimulation is provided by the inclusion of subset-specific cytokine markers, and proliferation markers. Development and optimisation of this panel was performed on freshly isolated cells from adult BALB/c lungs and small intestine lamina propria, and ex vivo stimulation with phorbol 12-myrisate 13-acetate, ionomycin and pro-ILC activating cytokines. O_TBL View this table: org.highwire.dtl.DTLVardef@1b1415forg.highwire.dtl.DTLVardef@3adc6forg.highwire.dtl.DTLVardef@5e0aeaorg.highwire.dtl.DTLVardef@1e9e61org.highwire.dtl.DTLVardef@2ff463_HPS_FORMAT_FIGEXP M_TBL O_FLOATNOTable 1.C_FLOATNO O_TABLECAPTIONSummary Table C_TABLECAPTION C_TBL Ethical compliance statementAll mouse experiments were performed in accordance with the recommendations in the Guide for the Use of Laboratory Animals of Imperial College London, with the ARRIVE (Animal Research: Reporting of In Vivo Experiments) guidelines. All animal procedures and care conformed strictly to the UK Home Office Guidelines under the Animals (Scientific Procedures) Act 1986, and the protocols were approved by the Home Office of Great Britain.