Potassium sulfate

M. marinum strain M (ATCC BAA-535) and ∆mmpL7 mutants (Cambier et al., 2014b (link)) expressing either TdTomato, Wasabi, or EBFP2 under the control of the msp12 promoter were grown under hygromycin (Thermo-Fisher) selection in 7H9 Middlebrook’s medium (Difco) supplemented with 10% OADC (Fisher), 0.2% glycerol, and 0.05% Tween-80 (Sigma). Where noted bacteria were also grown in glyceroL-alanine-salts (GAS) medium, recipe (%weight/volume) in 18 mM sodium hydroxide in Milli-Q water pH 6.6 +/- 0.05% Tween-80: 0.03% BactoCasitone (BD Science), 0.005% ferric ammonium citrate (Sigma), 0.4% potassium phosphate dibasic anhydrous (VWR), 0.2% citric acid, anhydrous (VWR), 0.1% L-alanine (Sigma), 0.12% magnesium chloride, heptahydrate (VWR), 0.06% potassium sulfate (VWR), 0.2% ammonium chloride (VWR), and 1% glycerol. To prepare heat-killed M. marinum, bacteria were incubated at 80°C for 20 min. To prepare fixed bacteria, bacteria were incubated in described concentrations of glutaraldehyde (Sigma) and/or paraformaldehyde (Thermo-Fisher) for 1 hr at 23°C, followed by three washes with PBS prior to experimental use.

RIT was obtained from AbbVie Deutschland GmbH & Co. KG (Ludwigshafen, Germany). The copolymer PVPVA (Kollidon^®® VA64) with a weight-average molar mass of 65,000 g/mol was purchased from BASF SE (Ludwigshafen, Germany). Hydrochloric acid (10%) was purchased from AppliChem GmbH (Darmstadt, Germany), and potassium acetate (purity ≥ 99%) was purchased from Carl Roth GmbH & Co. KG (Karlsruhe, Germany). Acetone (purity ≥ 99.8%), sodium dihydrogen phosphate (purity ≥ 99%), and disodium hydrogen phosphate dihydrate (purity ≥ 99.5%) were purchased from Merck KGaA (Darmstadt, Germany). Magnesium nitrate hexahydrate (purity ≥ 98%), potassium nitrate (purity ≥ 99%), and potassium sulfate (purity ≥ 99%) were purchased from VWR International GmbH (Darmstadt, Germany). All substances were used without further purification. Water was freshly prepared with a Milli-Q^®® Advantage A10 purification system from Merck KGaA (Darmstadt, Germany).

ABTS (2,2′-azinobis(3-ethylbenzothiazoline 6-sulfonic acid)), Trolox^® (6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid), DTT (dl-dithiothreitol), diatomaceous earth (Hyflo^® Super Cel^®), Trizma^® base, fluorescein sodium salt, and formic acid (reagent grade ≥ 95%) were purchased from Sigma-Aldrich (Steinheim, Germany). AAPH (2,2′-azobis(2-amidinopropane)) (Acros Organics), Tris (ultrapure), potassium dihydrogen phosphate, potassium sulfate, sodium phosphate dibasic, sodium chloride, ortho-boric acid, glycine (proteomics grade), and methanol (HPLC–MS grade) were provided by VWR International Eurolab S.L. (Barcelona, Spain). Glacial acetic acid, sulfuric acid, and sodium hydroxide were supplied by Fisher Scientific (Madrid, Spain). SDS (sodium dodecyl sulfate, purissimum-CODEX) and nitric acid (65% p/p) were obtained from Panreac (Barcelona, Spain). Acetonitrile (HPLC grade), acetone, glycerol, and bromophenol blue indicator (ACS reagent) were supplied by Merck (Darmstadt, Germany). Octadecyl C18 sorbent was from Phenomenex (Madrid, Spain), while absolute ethanol was from J.T. Baker (Deventer, The Netherlands). Anhydrous magnesium sulfate (99.5% min powder) was provided by Alfa Aesar (Karlsruhe, Germany). Deionized water (resistivity >18 MΩ/cm) was obtained through a Milli-Q SP^® Reagent Water System (Millipore Corporation Bedford, Bedford, MA, USA).