Streptomycin

Bacteria were subjected to 20 times the MIC₅₀ of fusidic acid (FA) (250 μg/ml; Sigma-Aldrich), nalidixic acid (330 μg/ml; Sigma-Aldrich), isoniazid (2 μg/ml; Sigma-Aldrich), linezolid (20 μg/ml; Sigma-Aldrich), streptomycin (20 μg/ml; SERVA), rifampicin (20 μg/ml; Sigma-Aldrich), Eis inhibitor 1a^* (10 μM) (Chen et al., 2012 (link); Green et al., 2018 (link)), or DMSO (solvent control; Merck Millipore) for 1 h.

Both free and fluorochrome conjugated antibodies (Abs): CD4 Pacific Blue, CD-80-FITC, CD-40-PEcy5, CD86-PE, MHC-II-Per-CPcy5-5, TIM-3-APC. All reagents used in cytokine ELISA experiment were procured from eBiosciences (San Diego, CA) and BD Pharamigen (San Diego, CA). Fetal Bovine Serum (FBS) was purchased from GIBCO (Grand Island, NY) and Biological Industries (Kibbutz Beit Haemek 25115, Israel). Penicillin, Streptomycin, L-glutamine, L-pyruvate and tissue culture grade plastic wares used in experiments were purchased from Serva (Heidelbergh, Germany) and BD Biosciences (Bedford, MA). Abs against pSTAT-1 and pSTAT-4 were procured from BD Biosciences (San Diego, CA). All other reagents and chemicals were purchased from Sigma Aldrich (St.Louis, MO) or otherwise mentioned.

NSC34 cells were used as motor neuronal cell model. Cells were routinely used in our lab^{11 (link),13 (link),30 ,31 (link)}. Briefly, cells were maintained in Dulbecco’s Modified Eagle Medium (DMEM) high glucose medium (Euroclone, Pero, MI, Italy) supplemented with 1 mM glutamine (Euroclone), penicillin (SERVA, Electrophoresis GmbH, Heidelberg, Germany) and streptomycin (SERVA) and 5% fetal bovine serum (Sigma-Aldrich). Cells were grown at 37 °C with 5% CO₂. To differentiate NSC34, retinoic acid 1 μM (Sigma-Aldrich) was added to full medium for 24 hrs before transfection and maintained for 72 hrs. C2C12 cells were used as muscle model. Cells were cultured as previously described^{32 (link)}, and maintained in DMEM high glucose medium completed with glutamine, penicillin, streptomycin and 10% of fetal bovine serum (GIBCO, Thermo Scientific Life Sciences Research, Waltham, MA, USA). Cells were grown at 37 °C with 5% CO₂. C2C12 were differentiated by supplementing medium with 2% Horse Serum (HS; Sigma-Aldrich), instead of 10% FBS, for 72 hrs.

α-T, H₂O₂, NADH, cytosine arabinoside were purchased in Sigma (Saint-Louis, MO, USA), K-252a, SL327, LY284002 and rottlerin were obtained from Calbiochem (San Diego, CA, USA), penicillin and streptomycin came from Serva (Heidelberg, Germany). The incubation media, Dulbecco’s modified Eagle Medium (DMEM) with l-glutamine, fetal calf blood serum were from Biolot Company (Saint-Petersburg, Russia). In the Immunoblotting section the information about various antibodies which were needed for experiments is given.

Five historical paper documents were offered for analysis by the Hellenic General State Archives (Athens, Greece). Four out of five documents dated back to 19th century (1840–1843), while the fifth one dated back to 20th century (1919). The samples were macroscopically examined and selected for further analysis, based on macroscopic patterns of biodeterioration such as discoloration, permanent staining, structural damage, and musty odor (Figure 1). Two sampling strategies were employed. In the first one, samples were collected from documents (surface area circa 30 cm²) demonstrating macroscopic biodeterioration patterns using sterile cotton swabs, whereas in the second one, sterile scalpels were used to remove small fragments (circa 0.5 cm²) directly from a heavily biodegraded area of documents. Both cotton swabs and paper fragments were kept at 4°C, in sterile vials till use. Cotton swabs were then used to inoculate Malt Yeast Extract agar plates (Lab M, UK) containing streptomycin (500 μg/ml) (Serva, Germany) for fungal or Nutrient Agar plates (Lab M, UK) for bacterial isolation. Agar plates were incubated at 30°C up to 7 days or up to 3 days, respectively. All fungal and bacterial isolates were kept at −80°C as glycerol stocks.

All cytokines and Abs used for ELISA and flowcytometry were procured from Becton Dickinson (Franklin Lakes, NJ), or otherwise mentioned. Growth media components were purchased from Hi-media (Mumbai, India) and Difco (Detroit, MI). Sodium acetate, lysozyme, proteinase K, RNase A, saponin, paraformaldehyde, CsA and other fine chemicals were purchased from SIGMA-ALDRICH (St. Louis, MO). CaeA was either procured from LKT Laboratories (St. Paul, MN) or purified from actinomycetes A. spitiensis. Aluminium sheets pre-coated with silica gel 60 F₂₅₄ were from Merck (Darmstadt, Germany). Fetal calf serum (FCS) was purchased from Harlan Sera Lab (Crawley Down, Great Britain). RPMI 1640, DMEM, HBSS and APO-BrdU TUNEL assay kit were bought from Invitrogen (Carlsbad, CA). L-glutamine, L-pyruvate, penicillin, concanavalin A and streptomycin were procured from Serva (Heidelberg, Germany). Thymidine-[methyl-³H] was the product of Amersham Pharmacia Biotech AB (Uppsala, Sweden).

The mouse motor-neuron-like hybrid cell line (NSC34), kindly provided by Dr. N.L. Cashman (University of British Columbia, Vancouver, CAN) [99 (link)], was maintained in DMEM medium supplemented with 5% fetal bovine serum (FBS)(F7524, Sigma-Aldrich), 1 mM L-glutamine (ECB3004D, Euroclone, Pero, Italy), and penicillin-streptomycin (penicillin, 31749.04, SERVA Electrophoresis GmbH, Heidelberg, Germany; streptomycin, 35500.01, SERVA Electrophoresis GmbH), at 37 °C in 5% CO₂. For experiments involving testosterone treatment, a dextran-treated charcoal-stripped FBS was used to selectively remove hormones [100 (link),101 (link)]. For the expression of the exogenous mutant proteins, NSC34 cells were transiently transfected with 0.7 μg of plasmid DNA using transferrin (T8150, Sigma-Aldrich) in conjunction with Lipofectamine reagent (18324012, Thermo Scientific Life Sciences Research, Waltham, MA, USA) to improve transfection efficiency.