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Arsenite

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Arsenite is a laboratory equipment used for the detection and quantification of arsenic compounds. It functions by converting arsenic compounds into a gaseous form, which can then be measured using spectroscopic techniques.

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Lab products found in correlation

Tunicamycin, by Merck Group (4 mentions) Fugene 6, by Promega (4 mentions) Cycloheximide (chx), by Merck Group (4 mentions) Thapsigargin, by Merck Group (3 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions) Las af software, by Leica Biosystems (3 mentions) Emetine, by Merck Group (2 mentions) Mg132, by Merck Group (2 mentions) Isrib, by Merck Group (2 mentions) Goat serum, by Vector Laboratories (2 mentions) Fetal calf serum (fcs), by Thermo Fisher Scientific (2 mentions) Nf200 antibody, by Merck Group (2 mentions) Dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (2 mentions) Dapi staining, by Vector Laboratories (2 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions) Glutamax, by Thermo Fisher Scientific (2 mentions) Paraformaldehyde (pfa), by Merck Group (2 mentions) Triton x 100, by Merck Group (2 mentions) Phosphate buffered saline (pbs), by Merck Group (2 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)

32 protocols using arsenite

1

Stress Granule Induction and Disassembly

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Unless otherwise stated SG were induced using arsenite (Sigma) at a final concentration of 0.1mM for 1h at 37°C except for MEFs (0.5mM for 1h) and the disassembly was forced by adding cycloheximide (Sigma) at 10μg/ml 30min after the arsenite treatment. P-eIF2α independent SG were induced using hippuristanol (Kind gift of J. Pelletier, McGill University) at 1μM for 1h. Phosphorylation of eIF2α was induced either by treatment with arsenite as described above, tunicamycin (Sigma) at 5μg/ml for 6h or by UV irradiation at 20mJ/cm2 using a Stratalinker 1800 (Stratagene). ISRIB (Sigma) was added to the cells at a final concentration ranging from 10 to 1000nM and A-92 (Axon Medchem) from 100 and 1000nM at t = 0hp.i and for the indicated times.
Brocard M., Iadevaia V., Klein P., Hall B., Lewis G., Lu J., Burke J., Willcocks M.M., Parker R., Goodfellow I.G., Ruggieri A, & Locker N. (2020). Norovirus infection results in eIF2α independent host translation shut-off and remodels the G3BP1 interactome evading stress granule formation. PLoS Pathogens, 16(1), e1008250.
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2

Permeabilized C. elegans Embryo Drug Treatment

Cited 1 time
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Drug treatment of C. elegans embryos was performed on permeabilized eggs. For this, young adult hermaphrodites were injected with perm-1 dsRNA and incubated at 20°C for 16 h after the injection. perm-1(RNAi) embryos were than extruded from the gravid hermaphrodite in a solution of egg buffer or 20 mM arsenite (MerckMillipore) diluted in egg buffer or 20 mM arsenite plus 250 μg/ml cycloheximide (Sigma Life Sciences, C7698-1G; Lee et al., 2020 (link)) diluted in egg buffer on a 22×40 mm coverslips. The coverslips with dissected worms and embryos were incubated for 1 h at room temperature in a humid chamber.
After the incubation time, the coverslip was mounted on a 3% agarose pad for imaging. Imaging was performed using the Leica DM6000 described above. Images were acquired using the 63×/1.4 numerical aperture (NA) objective and the LAS AF software (Leica Biosystems). The percentage of granule-containing embryos was counted.
Abbatemarco S., Bondaz A., Schwager F., Wang J., Hammell C.M, & Gotta M. (2021). PQN-59 and GTBP-1 contribute to stress granule formation but are not essential for their assembly in C. elegans embryos. Journal of Cell Science, 134(22), jcs258834.
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3

Temporal and Spatial Gene Expression Analysis in Drosophila

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For temporal and regional gene expression targeting experiments (TARGET), flies were grown at 18°C, collected over 1-2 days, and then reared in 29°C for up to 7 days before being sacrificed. For clonal analysis, clones were induced by heatshock and labeled with the mosaic analysis with repressible cell marker (MARCM) method. For these experiments, animals were reared at 25°C until eclosion, collected over 1 day, aged for 5 days, and then heatshocked at 37°C for 20 minutes in a Lauda circulating water bath. After heatshock, flies were reared at 25°C for 10 or 60 days.
For ex vivo insulin assays, intestines from animals starved for 4-days were incubated in Grace’s insect media supplemented with 200-1600 nM human insulin (Sigma) for 10 minutes and then fixed. For ex vivo arsenite treatment, intestines from females aged for 8-10 days on normal diet were incubated in Krebs-Ringer media supplemented with 1mM arsenite (Sigma) for 60 minutes and then fixed.
Luhur A., Buddika K., Ariyapala I.S., Chen S, & Sokol N. (2017). Opposing post-transcriptional control of InR by FMRP and LIN-28 adjusts stem cell based tissue growth. Cell reports, 21(10), 2671-2677.
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4

Beas-2B Cell Culture and Treatment

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BEAS-2B (ATCC, Manassas, VA), was cultured in BEGM media (Lonza, Walkersville, MD), with or without 5% fetal bovine serum (FBS) for 8 weeks. All studies were performed using BEAS-2B cells between passage 10 and passage 30, with passage 1 defined as the thawed cells from the supplier. Trypsin-EDTA (0.25%) was used to remove cells from culture flasks for sub-culturing. Cell cultures were incubated in humidified 95% air: 5% CO2. Two million cells were seeded in 75 cm2 culture flasks and sub-cultured at 90% confluence. BEAS-2B cells referred to in this study as “control” were cultured for appropriate passage-matched durations in BEGM without FBS and without arsenite. Cells referred to as “FBS-exposed” were cultured in BEGM, with 5% FBS, without arsenite, for 8 weeks. Cells referred to as “arsenite-exposed” were cultured in BEGM, without FBS, with 1 μM arsenite (Sigma, St. Louis, MO), for 8 weeks. The identity of the BEAS-2B cell line was confirmed using commercial forensic DNA testing based on polymorphic STR markers. BEAS-2B cells were assayed for mycoplasma contamination every two weeks using the MycoAlert system (Lonza, Switzerland). No indication of mycoplasma contamination was found during the course of this study.
Zhao F, & Klimecki W.T. (2014). CULTURE CONDITIONS PROFOUNDLY IMPACT PHENOTYPE IN BEAS-2B, A HUMAN PULMONARY EPITHELIAL MODEL. Journal of applied toxicology : JAT, 35(8), 945-951.
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5

Cell Culture and Transfection Techniques

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HeLa cells, HEK293T cells and U2OS cells were maintained in DMEM (Life Technologies) supplemented with 1% penicillin/streptomycin and 10% fetal bovine serum (Hyclone). Cells were transfected with 1 µg of total DNA per 4 × 105 cells, unless indicated otherwise, using JetPrime (PolyPlus transfections) according to the manufacturer's instructions. If more than one plasmid was used to transfect cells, the amount of each plasmid used per transfection reaction was constant. Twenty-four hours after transfection, cells were fixed or lysed. For siRNA transfection, 20 nM of siRNA was used to transfect 150,000 cells using Lipofectamine 2000 (Invitrogen) reagent according to manufacturer's instructions. Cells were treated with 500 mM arsenite (Sigma-Aldrich) for 1 h and with 1 µM Emetine (Sigma-Aldrich) for 50 min (Cinti et al. 2017 (link)).
Rao S., Cinti A., Temzi A., Amorim R., You J.C, & Mouland A.J. (2018). HIV-1 NC-induced stress granule assembly and translation arrest are inhibited by the dsRNA binding protein Staufen1. RNA, 24(2), 219-236.
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6

Cellular stress response assay

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Cells were seeded 48 h prior to stress treatment. Cells were treated with 5 µg/mL anisomycin (Calbiochem), 100 µM arsenite (Sigma), or 3 mM DTT (Promega) for 3 h and harvested in ice-cold PBS.
Hollerer I., Curk T., Haase B., Benes V., Hauer C., Neu-Yilik G., Bhuvanagiri M., Hentze M.W, & Kulozik A.E. (2016). The differential expression of alternatively polyadenylated transcripts is a common stress-induced response mechanism that modulates mammalian mRNA expression in a quantitative and qualitative fashion. RNA, 22(9), 1441-1453.
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7

Cell Lines and Stress Granule Induction

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HeLa (CCL-2), HEK293T (CRL-3216) and RAW 264.7 (TIB-71) cell lines were obtained from ATCC and were grown in DMEM supplemented with 10% of foetal calf serum. We have not authenticated these ATCC cell lines. Immortalised bone marrow-derived macrophages49 (link) cell line originally derived from C57BL/6J mice were provided by Thomas Henry (CIRI, Lyon, France) maintained in DMEM supplemented with 10% FCS and 10% spent medium from L929 cells that supply MC-CSF. All cells were routinely tested and were mycoplasma free. When indicated, cells were treated with 0.5 mM arsenite (Sigma) for 30 min to induce stress granules and P-bodies and treated with 5 μM thapsigargin (Sigma) 4 h to induce U-bodies.
Louche A., Blanco A., Lacerda T.L., Cancade-Veyre L., Lionnet C., Bergé C., Rolando M., Lembo F., Borg J.P., Buchrieser C., Nagahama M., Gérard F.C., Gorvel J.P., Gueguen-Chaignon V., Terradot L, & Salcedo S.P. (2023). Brucella effectors NyxA and NyxB target SENP3 to modulate the subcellular localisation of nucleolar proteins. Nature Communications, 14, 102.
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8

Antibody Characterization for Neurodegenerative Research

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Mouse monoclonal anti-ubiquitin, HuR, and GFP were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Mouse monoclonal anti-α tubulin was from Invitrogen. Mouse monoclonal Anti-βIII Tubulin (2G10), p62/SQSTM1, MAP2 and FMRP was obtained from Millipore (Billerica, MA). Rabbit polyclonal anti-N-terminus FUS was from Bethyl Labs (Montgomery, TX, USA). Anti-TDP-43 and G3BP antibodies were from Proteintech (Chicago, IL, USA). Rabbit polyclonal anti-GFP antibody was from Rockland Immunochemicals Inc. (Gilbertsville, PA, USA). Mouse monoclonal anti-GFP antibody was from Santa Cruz Biotechnology, Inc (Dallas, TX, USA).Mouse Anti-Ataxin-2 was purchased from BD Biosciences (Sparks, MD, USA). Rabbit polyclonal anti-GAPDH antibodies were from Cell Signaling Technology (Beverly, MA, USA). Propidium iodide was purchased from Dojindo (Kumamoto, Japan). Emetine, and Arsenite was purchased from Sigma-Aldrich.
Mitsuhashi K., Ito D., Mashima K., Oyama M., Takahashi S, & Suzuki N. (2017). De novo design of RNA-binding proteins with a prion-like domain related to ALS/FTD proteinopathies. Scientific Reports, 7, 16871.
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9

Induction of Stress Granules in Myogenic and Fibroblast Cells

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C2C12 cells (American Type Culture Collection/Cederlane) were maintained in growth medium (DMEM, 10% fetal bovine serum [Wisent Bioproducts, St-Bruno, Canada], 100 U/ml penicillin, and 100 μg/ml streptomycin). To induce myogenic differentiation, C2C12 cells were allowed to become confluent on Matrigel-coated (BD Biosciences/Fisher Scientific) plates, and the medium was switched to differentiation medium (DMEM, 2% horse serum [PAA Laboratories, Piscataway, NJ], 100 U/ml penicillin, and 100 μg/ml streptomycin). Control (GM01653, GM03377, GM03523) and DM1 (GM03991, GM03987, GM03132) human fibroblasts (Coriell Cell Repositories, Coriell Institute for Medical Research, Camden, NJ; Table 1) were grown according to instructions.
Induction of SG formation was done by treating cells with 0.5 mM arsenite (Sigma-Aldrich, Oakville, Canada) for 45 min at 37°C. Alternatively, SG formation was also induced using 1 μM thapsigargin (Sigma-Aldrich) for 60 min at 37°C or by heat shock for 45 min at 45°C. When specified, C2C12 cells were treated with CHX (50 μg/ml; Sigma-Aldrich) or puromycin (Puro, 10 μg/ml; Wisent Bioproducts) just before and for the entire duration of the arsenite treatment.
Ravel-Chapuis A., Klein Gunnewiek A., Bélanger G., Crawford Parks T.E., Côté J, & Jasmin B.J. (2016). Staufen1 impairs stress granule formation in skeletal muscle cells from myotonic dystrophy type 1 patients. Molecular Biology of the Cell, 27(11), 1728-1739.
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10

Immunofluorescence Assay for ER Stress

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Mouse monoclonal anti-GADD 153 (B-3) antibody (Santa Cruz Biotechnology, Dallas, TX, USA); polyclonal antibody against neuronal class III β-tubulin (Covance, Princeton, NJ, USA); monoclonal anti-CtBP2 antibody (BD Biosciences, San Jose, CA, USA); polyclonal antibody against p-eIF2α (Cell Signaling, Danvers, MA, USA); polyclonal anti-myosin 7a antibody (Proteus Biosciences, Ramona, CA, USA); secondary goat anti-rabbit antibody conjugated with Texas Red (Abcam, Cambridge, MA, USA); rhodamine phalloidin, Invitrogen (Life Technologies, Carlsbad, CA, USA); HEK293 cells (Innoprot, Biscay, Spain); geneticin, gentamicin, tunicamycin, cycloheximide, arsenite, saponin and HEK aph(3′) cells (Sigma Aldrich, St. Louis, MO, USA); hygromycin (PAA Laboratories, Cansera, Canada); nucleotide primers (Microsynth, Balgach, Switzerland); cell culture media and trypsin (Life Technologies).
Oishi N., Duscha S., Boukari H., Meyer M., Xie J., Wei G., Schrepfer T., Roschitzki B., Boettger E.C, & Schacht J. (2015). XBP1 mitigates aminoglycoside-induced endoplasmic reticulum stress and neuronal cell death. Cell Death & Disease, 6(5), e1763-.
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