Fc block solution

For multicolor flow cytometry analysis whole blood and bone marrow (BM) of the mice was collected. Erythrocytes in 100µl whole blood were lysed using a TQ-Prep Workstation (Beckmann Coulter, Brea, CA, USA), for analysis of BM, 1x10⁵ cells/stain were used. Cells (lysed blood cells and BM) were resuspended in 50µl Fc-block solution (eBioscience, San Diego, CA, USA) and incubated for 10min at room temperature. Staining for antibody panels was then carried out at 4°C for 30min as described previously (39 (link)). Cells were analyzed using a CytoFLEX S flow cytometer and data was analyzed with the help of the Kaluza analysis software (both: Beckman Coulter, Brea, CA, USA).

For flow cytometry analysis whole blood and bone marrow (BM) of animals was collected. Erythrocytes in 100 µL whole blood were lysed by 0.12% formic acid for 10 s, followed by a neutralization step and subsequent fixation in 4% paraformaldehyde. For analysis of BM, 1 × 10⁵ cells/stain and for blood samples 35 µL of whole blood/stain was used. For ex vivo experiments, cells were collected 24 h after radon or mock exposure. Cells (lysed blood cells, BM as well as monocytes and macrophages) were resuspended in 50 µL Fc-block solution (eBioscience, San Diego, CA, USA) and incubated for 10 min at room temperature. Staining for antibody panels was then carried out at 4 °C for 30 min as described previously [33 (link)] and as it can be seen in Table A1 in Appendix A. For 2′,7′-Dichlordihydrofluorescein-diacetat (DCF) Assay, cells were washed and treated with either 2 µM DCF or 0µM DCF (Invitrogen, Waltham, MA, USA) for 90 min at standard culture conditions in the dark. Subsequently, cells were detached and analyzed in a flow cytometer. For evaluation, ∆mean fluorescence intensity (MFI) of (2 µM DCF)–(0 µM DCF) was calculated. Cells were analyzed using a CytoFLEX S flow cytometer and data was analyzed with the help of the Kaluza analysis software (both: Beckman Coulter, Brea, CA, USA). Gating strategies are depicted in Appendix A Figure A1 and Figure A2.