2 2 azinobis 3 ethylbenzothiazoline 6 sulfonic acid (abts)

Manufactured by Roche

Sourced in Germany, United States, Switzerland, Japan

ABTS is a laboratory reagent used in colorimetric assays. It is a substrate for various enzymes, including peroxidases, which catalyze its oxidation to produce a colored product that can be measured spectrophotometrically.

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Lab products found in correlation

Potassium persulfate, by Merck Group (5 mentions) Folin ciocalteu reagent, by Merck Group (3 mentions) Maxisorp, by Thermo Fisher Scientific (2 mentions) Polyclonal goat igg anti human kappa, by Southern Biotech (2 mentions) Biotinylated anti ifnγ detection antibody, by BD (2 mentions) Bovine serum albumin (bsa), by Merck Group (2 mentions) Yeast transformation kit, by Merck Group (1 mentions) S cerevisiae bj5465 strain, by LGC (1 mentions) Zymoprep yeast plasmid miniprep 2, by Zymo Research (1 mentions) Qiaquick gel extraction kit, by Qiagen (1 mentions) High pure plasmid isolation kit, by Roche (1 mentions) Deuterated solvent, by Merck Group (1 mentions) 3 4 5 dimethyl 2 thiazolyl 2 5 diphenyl 2h tetrazolium bromide, by Merck Group (1 mentions) Methyl thiazolyl tetrazolium (mtt), by Merck Group (1 mentions) Hrp conjugated anti flag m2 antibody, by Merck Group (1 mentions) Hrp conjugated goat anti rabbit antibody, by Bio-Rad (1 mentions) Nunlock microplates, by Greiner (1 mentions) Hydrochloric acid (hcl), by Avantor (1 mentions) 6 hydroxy 2 5 7 8 tetramethylchromane 2 carboxylic acid trolox, by Merck Group (1 mentions) Ferulic acid, by Merck Group (1 mentions)

46 protocols using 2 2 azinobis 3 ethylbenzothiazoline 6 sulfonic acid (abts)

Maize Detoxification Mechanisms Examined

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Caryopses of Zea mays cultivar Cassila (KWS, Germany) were hydroponically grown for 7 days as described in Schulz et al. (2016) (link). For the analysis of detoxification products, the seedlings were incubated with the BOA-isomers for 24h (3 seedlings/ 20 ml of 0.5 mM of one of the isomers in tap water). For analysis of BOA-6-O-glc accumulation during short term exposure, seedlings were incubated 0, 10, 20, 30, 40, 50 and 60 min with 0.5 µM BOA-6-OH.
Analysis and determination of the detoxification products was performed by HPLC using the system and the method described below. For preparation of 500 ml incubation solution the isomers were pre-solved in 1 ml methanol and sonicated for 5 min before adding tap water.
Root extracts were performed as described in Schulz and Wieland (1998) . Peroxidase activity at living root surfaces was detected by bathing the roots (1 seedling / 15 ml Falcon tube) in 10 ml 0.1 mM acetate buffer pH 5.0 supplemented with 1.6 ml 2 mM ABTS (Roche Diagnostics, Germany) and 0.6 ml 15 mM H2O2 (Sigma-Merck, Germany). Sites with peroxidase activity immediately turned dark or black-green. Catalase activity elicited by BOA-6,5,4-OH was concluded from bubble formation according to Schellhorn and Stones (1992) (link) and Wheelis (2008) , presenting a non-destructive method.

Schulz M., Laschke L., Schütz V., Schackow O., Sicker D., Hofmann D., & Dörmann P. (2021). Survival of Plants During Short-Term Boa-Oh Exposure: Ros Related Gene Expression and Detoxification Reactions are Accompaniedwithfast Membrane Lipid Repair in Root Tips.

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Yeast Transformation Protocol for Protein Expression

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Yeast Transformation Kit was purchased from Sigma-Aldrich (St. Louis, MO, USA). High Pure Plasmid Isolation Kit and ABTS (2,2′azinobis (3ethylbenzothiazoline- 6 sulphonic acid)) were obtained from ROCHE. Phusion High-Fidelity DNA polymerase and Restriction enzymes (NotI and BamHI) were obtained from New England Biolabs. QIAquick gel extraction kit from Qiagen (Hilden, Germany)). Zymoprep™ Yeast Plasmid Miniprep II was purchased from Zymo Research (Tustin, CA, USA). S. cerevisiae BJ5465 strain was purchased from LGC Promochem (Barcelona, Spain). The α-factor preproleader (α_nat) was obtained from the pPICZα family of plasmids of Invitrogen (Waltham, MA, USA).

Aza P., de Salas F., Molpeceres G., Rodríguez-Escribano D., de la Fuente I, & Camarero S. (2021). Protein Engineering Approaches to Enhance Fungal Laccase Production in S. cerevisiae. International Journal of Molecular Sciences, 22(3), 1157.

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Measurement of LIF and LIFR Antibodies

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Mice were bled via facial vein after receiving the last booster. Serums were tested for production of LIF- and LIFR-specific antibodies in an ELISA. Briefly, 96-well plates were coated at 4 °C overnight with 200 ng/well of rtLIF and rtLIFR in a coating buffer (100 mM Na₂CO₃, 50 mM NaHCO₃ and pH 9.6). The plates were incubated with 1:200, 1:400, 1:800, 1:1,600, 1:3,200, 1:6,400 dilutions of the immune serums in TEN-TC buffer (50 mM Tris, 12 mM EDTA, 1.5 M NaCl, 0.5% Tween 20 and 2% casein) for 1 h at 37 °C. Washing with TEN-T buffer (TEN-TC without casein) was followed by incubation with polyclonal goat anti-mouse-IgG HRP-conjugated antibody (Invitrogen) diluted at ratio of 1:1,000 in TEN-TC buffer for 1 h at 37 °C. ABTS (Roche) and H₂O₂ was added and incubated for 15 min in darkness. In the end, the absorbance was measured at 405 nm by an ELISA instrument.

Ghanei Z., Mehri N., Jamshidizad A., Joupari M.D, & Shamsara M. (2020). Immunization against leukemia inhibitory factor and its receptor suppresses tumor formation of breast cancer initiating cells in BALB/c mouse. Scientific Reports, 10, 11465.

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Antioxidant Screening of Teucrium chamaedrys L.

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All of the solvents and reagents used for assessing antioxidant screening were purchased from Sigma-Aldrich Chemie (Buchs, Switzerland) except ABTS, which was bought from Roche Diagnostics (Roche Diagnostics, Mannheim, Germany). Cell culture medium and reagents for cytotoxicity testing were purchased from Invitrogen (Paisley, UK); MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide] was bought from Sigma-Aldrich Chemie. Deuterated solvents and internal standard for NMR-based metabolic profiling analyses were purchased from Sigma-Aldrich Chemie.
Leaves belonging to the species T. chamaedrys L. subsp. chamaedrys, a small eurimediterranean shrub characteristic of the coastal macchia vegetation [21 (link)], were collected at the “Castel Volturno” Nature Reserve (Caserta, Italy). Voucher specimens (CE0037) have been deposited at the Herbarium of the Department of Environmental, Biological and Pharmaceutical Sciences and Technologies, University of Campania “Luigi Vanvitelli”.

Piccolella S., Scognamiglio M., D’Abrosca B., Esposito A., Fiorentino A, & Pacifico S. (2021). Chemical Fractionation Joint to In-Mixture NMR Analysis for Avoiding the Hepatotoxicity of Teucrium chamaedrys L. subsp. chamaedrys. Biomolecules, 11(5), 690.

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Quantitative Analysis of scFv Binding to Immobilized Pol II

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A 96-well microtiter plate was coated with 0.3 μg of pol II per well overnight at 4°C. BSA was used as negative control. Solution was removed and wells were blocked with 150 μl of 5% milk in PBS for 1 hour at room temperature. Blocking solution was removed and 50 μl of 0–10 μM purified scFv were added, followed by incubation for 30 minutes at room temperature. Solution was removed and wells were washed with PBST, followed by 50 μl HRP-conjugated anti-Flag M2 antibody (Sigma, A8592–1MG) diluted 1:1000 in 5% milk PBS and incubation for 30 minutes at room temperature. Solution was removed, wells were washed four times with PBST, 50 ul per well of 100 ug/ml ABTS (2,2’-azino- bis(3-ethylbenzthiazoline-6-sulphonic acid) (Roche, Cat # 11684302001) in 0.1 M citrate buffer pH 4.0 containing 0.01 % hydrogen peroxide were added, and A₄₀₅ was recorded.

Jagota M., Townshend R.J., Kang L.W., Bushnell D.A., Dror R.O., Kornberg R.D, & Azubel M. (2021). Gold nanoparticles and tilt pairs to assess protein flexibility by cryo-electron microscopy. Ultramicroscopy, 227, 113302.

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Quantifying Antigen-Antibody Interactions

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2HB 96-well plates were coated with 100 μL/well of rCD99 antigens diluted at 5 μg/mL in carbonate buffer pH 9.6 and kept at 37°C for 16 to 18 hours. After five washes in PBS containing Tween-20 0.05%, the plates were blocked with BSA (#A4503, Sigma) 1% in PBS (150 μL/well) and kept for 1 hour at 37°C. After washing, serial dilutions of dAbd C7 (Diatheva Srl) in PBS-BSA were added to the wells and the plates incubated for 90 minutes at 37°C. After washing, an anti-scFv polyclonal antibody (Diatheva Srl) was added and kept for 1 hour at 37°C. The immunoreactive signals were highlighted after further addition of a goat anti-rabbit HRP-conjugated antibody (#1706515, Bio-Rad, RRID: AB11125142). After incubation and washing, the substrate ABTS (#11684302001, Roche Diagnostic) was added, and the absorbance values were obtained reading at 405 nm with a microplate reader (Glomax Multi Detection System).

Manara M.C., Manferdini C., Cristalli C., Carrabotta M., Santi S., De Feo A., Caldoni G., Pasello M., Landuzzi L., Lollini P.L., Salamanna F., Dominici S., Fiori V., Magnani M., Lisignoli G, & Scotlandi K. (2023). Engagement of CD99 Activates Distinct Programs in Ewing Sarcoma and Macrophages. Cancer Immunology Research, 12(2), 247-260.

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ELISA for IFN-γ and IL10 Quantification

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IFN-γ and IL10 in culture supernatants were monitored by ELISA by binding to the solid phase anti-IFN-γ and IL10 capture antibody (each 1 µg/mL), respectively, and detection by the biotinylated anti-IFN-γ or anti-IL10 detection antibody (0.5 µg/mL), respectively. The reaction product was visualized by a peroxidase-streptavidin conjugate (1:10,000) and ABTS (Roche Diagnostics, Basel, Switzerland).

Hombach A.A., Geumann U., Günther C., Hermann F.G, & Abken H. (2020). IL7-IL12 Engineered Mesenchymal Stem Cells (MSCs) Improve A CAR T Cell Attack Against Colorectal Cancer Cells. Cells, 9(4), 873.

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Quantifying Antibody Responses to Ovalbumin

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Total IgE was determined by sandwich ELISA, using a commercial test (Cat. No. 157718; Abcam, Cambridge, UK) following the manufacturer’s protocol. Specific to OVA, serum IgA and IgG and feces IgA were measured by indirect ELISA (16). The antigen (1 µg/well in 10 mM PBS, pH 7.4) was coated onto 400 Nunlock microplates (Greiner Bio-One Gmbh, Frickenhausen, Germany) and incubated at 37 °C for 1 h. Next, the plates were blocked with 1.5% gelatin diluted in PBS and incubated under the same conditions. Following blocking, the plates were washed with PBST (PBS + 0.05% Tween 20). Serial dilutions of serum and feces extracts (50 µL) were added to the plates and incubated for 1 h at 37 °C. After a washing step, the plates were incubated with HRP-labeled antibodies specific to mouse IgG (1:1000) or IgA (1:1000) for 1 h at 37 °C. The plates were washed and incubated for 1 h at RT with ABTS (Roche Diagnostics GmbH, Mannheim, Germany). The absorbance was measured at 405 nm on a Jupiter UVM spectrophotometer (ASYS-Hitech GmbH, Eugendorf, Austria). The endpoint titers (EPTs) were expressed as the reciprocal dilution of the last sample dilution of 0.1 OD above the negative control [42 (link),43 (link),44 (link),45 (link)].

Fuc E., Złotkowska D., Wasilewska E, & Wróblewska B. (2021). OVA-Experienced CD4+ T Cell Transfer and Chicken Protein Challenge Affect the Immune Response to OVA in a Murine Model. International Journal of Molecular Sciences, 22(12), 6573.

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Monoclonal Antibody Generation and CII Antibody Responses

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Several monoclonal antibodies were generated from bCXI-immunized mice and their binding specificity was determined. Whole blood was collected from the mouse retro-orbital venous sinus on day 21 and day 70 after immunization with CII. 32 sera samples in total from each time point were measured for the anti-CII and anti-CXI antibody responses. 96-well flat-bottom ELISA plates (Nunc MaxiSorp; Denmark) were coated overnight with 5 µg/ml of purified rCXI, rCII, and bCXI, either native or denatured, or with 5 µg/ml synthetic peptides in PBS at 4°C. After the blockage with 3% non-fat milk in PBS at room temperature (RT) for 2 h, purified antibodies diluted according to a previously determined concentration and 1:1,000 diluted serum samples were incubated at RT for 2 h. The antibody titers were evaluated using HRP-conjugated anti-kappa-specific antibody (1:4,000, Southern Biotech) and ABTS (Roche) as detect system. For isotype specific assessment, biotinylated goat anti-mouse-IgM, -IgG1, -IgG2a, -IgG2b, or -IgG3 reagents (Southern Biotech) were used.

Tong D., Lönnblom E., Yau A.C., Nandakumar K.S., Liang B., Ge C., Viljanen J., Li L., Bãlan M., Klareskog L., Chagin A.S., Gjertsson I., Kihlberg J., Zhao M, & Holmdahl R. (2018). A Shared Epitope of Collagen Type XI and Type II Is Recognized by Pathogenic Antibodies in Mice and Humans with Arthritis. Frontiers in Immunology, 9, 451.

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Eculizumab-Based Quantification of C5 in PNH

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Eculizumab (50 μl of a 2 μg/ml solution) was coated onto 96-well microtiter plates (MaxiSorp; Nunc) as described previously (19 (link)). After washing, the plate was blocked with a 1% solution of BSA in PBS. Then, wells were exposed to a 1:200 dilution of a serum sample (50 μl). The samples consisted of PNH patient sera alone or mixed with NHS in ratios as specified. After washing, the C5 bound to the coated Eculizumab was detected by sequentially applying two detection antibodies (with intermediary washing): a polyclonal goat anti-human C5 antibody (Quidel), followed by a HRP-coupled donkey-anti-goat antibody, followed by color development with ABTS (Roche) in the presence of 0.03% H₂O₂. Absorbance was quantified at 405 nm.

Harder M.J., Höchsmann B., Dopler A., Anliker M., Weinstock C., Skerra A., Simmet T., Schrezenmeier H, & Schmidt C.Q. (2019). Different Levels of Incomplete Terminal Pathway Inhibition by Eculizumab and the Clinical Response of PNH Patients. Frontiers in Immunology, 10, 1639.

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