Ethylene diamine tetraacetic acid (edta)

Cardiac stromal cells were incubated for 15 min at 4 °C in blocking buffer (DPBS (SIGMA) with 4% HIFCS, 3% mouse serum (SIGMA), 3% rat serum (SIGMA), and 2 mM EDTA (Euromedex, Souffelweyersheim, France)) with 5 µg/mL anti CD16/CD32 (Biolegend, Ozyme) as previously described [31 (link)]. Cells were then incubated on ice with conjugated monoclonal antibodies and diluted at their optimal concentrations (Supplementary Table S2) in FACS buffer (DPBS with 4% HIFCS, 2 mM EDTA) for 30 min (FACS) or 45 min (cell sorting). Necrotic cells were excluded by staining with Live/Dead Aqua or Live Dead Yellow fluorescent reactive dyes (Life Technologies, Thermo Fisher Scientific).
Cardiac stromal cell subsets were isolated by high-speed sorting with the BD InfluxTM cell sorter (BD Biosciences, Allschwil, Switzerland) and cMSCs were cell sorted based on CD45^– CD31^– Sca-1⁺ CD140a⁺ expression, macrophages on CD45⁺ CD64⁺ MHCII⁺ and vascular ECs were cell sorted on CD45^– CD140a^– CD31⁺ Sca-1⁺ expression (Supplementary Table S2).

All binding experiments were performed in TBS buffer (20 mM Tris, 125 mM NaCl, 2 mM CaCl₂) containing 0.5 mg/mL BSA (Sigma). Non-specific binding to M. tuberculosis was prevented by incubating the bacteria for 30 min at room temperature in TBS buffer containing 2 mg/mL BSA. Bacteria were then incubated with native CL-LK (5 μg/mL, prepared as in [17 (link)]) at 37°C for 1 h in the presence or absence of 20 mM EDTA (Euromedex) or 50 mg/mL purified mannan. After washing in TBS, bacteria were incubated with a biotinylated monoclonal anti-CL-K1 antibody (HYB14-29 [18 (link)]) at 2 μg/ml for 1 h at room temperature. Secondary detection was achieved using APC-coupled Streptavidin (eBioscience) (for 20min at room temperature). Bacteria were then washed and fixed for 2 h at room temperature in PBS containing 4% paraformaldehyde (Polyscience)