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2 protocols using isoniazid

1

Teratogens Screening in Zebrafish

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Test compounds used in this study are listed in Table 1. These tested compounds are known to be teratogens inducing cleft palate in mammals and have been classified into various categories as a result of being tested in zebrafish experiments or chemical safety assays (Hillegass et al., 2008 (link); Selderslaghs et al., 2009 (link); Ito and Handa, 2012 (link); Lee et al., 2012 (link); Teixido et al., 2013 (link); Yamashita et al., 2014 (link); Inoue et al., 2016 (link); Martinez et al., 2018 (link); Cassar et al., 2019 (link)). The test compounds and exposure concentrations were determined based on Liu et al., 2020 (link). The exposure concentrations were as follows: hydroxyurea (1 mM, Sigma-Aldrich), valproic acid (7.5–30 μM, Wako), salicylic acid (100–400 μM, Wako), boric acid (1 mM, Wako), and caffeine (0.5–2 mM, Wako), which were diluted from stock solutions prepared with distilled water (Life Technologies), and imatinib (250 μM, Tokyo Chemical Industry), retinoic acid (10–50 nM, Tokyo Chemical Industry), thalidomide (400 μM, Tocris Bioscience), methotrexate (50–200 μM, Wako), warfarin (15–60 μM, Wako), phenytoin (1 mM, Wako), dexamethasone (1 mM, Wako), 5-fluorouracil (1 mM, Wako), and isoniazid (1 mM, LKT Laboratories), which were diluted from stock solutions prepared with dimethyl sulfoxide (DMSO, Wako).
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2

Mycobacterium vanbaalenii Growth Inhibition

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Mycobacterium vanbaalenii (NRRL B-24157 strain, RIKEN BioResource Center, Tsukuba, Japan) was grown overnight at 28°C in 3.7% brain-heart infusion broth (Sigma, St. Louise, MO, USA). Cultures were then diluted 6-fold with broth containing the candidate compounds in 96-well, flat-bottom clear plates. Each well was inoculated with 0.200 mL of culture. Isoniazid (LKT laboratories, St. Paul, MN, USA) and 0.3% dimethyl sulfoxide (DMSO) were used as positive and negative controls, respectively. The plates were incubated at 28°C for 18 h, after which cell cultures were subjected to growth inhibition assays. Inhibition of bacterial growth was determined by measuring OD 595 with a Bio-Rad Model 680 microplate reader (Bio-Rad, Hercules, CA, USA).
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