One step tmb

Manufactured by Thermo Fisher Scientific

One-Step TMB is a ready-to-use substrate solution for the detection of horseradish peroxidase (HRP) in enzyme-linked immunosorbent assay (ELISA) applications. It provides a colorimetric readout that can be measured spectrophotometrically.

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Lab products found in correlation

High binding elisa plate, by Corning (2 mentions) Stop solution, by Thermo Fisher Scientific (2 mentions) Envision 2102 multilabel reader, by PerkinElmer (1 mentions) Superblock, by Thermo Fisher Scientific (1 mentions) Pmsf protease inhibitor, by Thermo Fisher Scientific (1 mentions) Kpl wash buffer, by LGC (1 mentions)

Close spelling variants of this product

One-Step Ultra TMB (10 citations) One-Step Ultra TMB ELISA HRP substrate solution (6 citations) One-step Ultra TMB substrate (6 citations) One-step TMB substrate (3 citations) One-step™ Ultra TMB-ELISA Substrate Solution (3 citations) One-Step Ultra TMB-ELISA (2 citations) One-step Turbo TMB (2 citations) One-step Ultra TMB-Blotting Solution (2 citations)

4 protocols using one step tmb

Quantifying Tau Protein Binding Profiles

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Example 5

Binding to recombinant WT (2N4R; SEQ ID NO:31) tau was analyzed by ELISA where full-length Tau protein (1 ng/mL or 10 ng/mL) was directly coated to the plate and incubated with different concentrations of either recombinantly- or hybridoma produced PT82 antibody (FIG. 2). After incubation with antibodies, plates were again washed and 50 μL per well of HRPO labelled anti-mouse antibody (GE Healthcare) (diluted 1:10000 in blocking buffer) was added. After another washing step detection was performed with “One step” TMB (Thermo Scientific) according to the manufacturers' instructions. Plates were analysed in EnVision® 2102 Multilabel Reader (Perkin Elmer, Waltham, Mass., USA). Binding curves were generated using GraphPad Prism7.0 software. As expected, lower coating concentrations of Tau resulted in lower maximal values (e.g. compare red to green curves where binding of recombinant antibodies to respectively 1 ng/mL or 10 ng/ml are shown). No substantial difference has been observed between binding profiles of recombinant and hybridoma produced antibodies.

US10633435B2. Anti-PHF-tau antibodies and uses thereof (2020-04-28). JANSSEN PHARMACEUTICA NV [US]. Inventors: Kristof Van Kolen [BE], Marc Mercken [BE], Linda Barone [US], Eilyn R. Lacy [US], Rupesh Nanjunda [US], John Wheeler [US], Jinquan Luo [US].

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Evaluating dTIT7 Binding to L-MC16

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To assess dTIT7 binding, wells of Sulfhydryl-BIND Surface Maleimide plates (Corning) were coated 20 hours with wild type (WT) and mutant L-MC16 in water at 4°C. After washing with PBS containing 0.02% Tween 20 (PBST), wells were blocked 1 hour with 10% superblock (Thermo) in PBST at room temperature. Biotinylated dTIT7 (10 μg dissolved in 10% superblock in PBST (1 mL) prepared as above was added to each well at 100 μl/well and incubated 30 min at room temperature for. After three PBST washes, 100 μl streptavidin-peroxidase (0.2 μg/ml) in 10% superblock in PBST containing 2% bovine serum albumin was added to each well and incubated 30 min. After three PBST washes, 100 μL of the peroxidase substrate one-step-TMB (Thermo) was added and incubated until the color developed. The reaction was stopped by adding 100 μl 2N sulfuric acid, and absorbance at 450 nm monitored using an ELISA plate reader.

Nonaka M., Mabashi-Asazuma H., Jarvis D.L., Yamasaki K., Akama T.O., Nagaoka M., Sasai T., Kimura-Takagi I., Suwa Y., Yaegashi T., Huang C.T., Nishizawa-Harada C, & Fukuda M.N. (2021). Development of an orally-administrable tumor vasculature-targeting therapeutic using annexin A1-binding D-peptides. PLoS ONE, 16(1), e0241157.

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Cytokine Quantification by ELISA

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IFNγ, IL-4, and IL-13 capture ELISAs were performed according to the manufacturer’s instructions (R&D Systems, Minneapolis, MN, USA). Briefly, the capture antibody was coated onto a high-binding ELISA plate (Corning) overnight. The following day, wells were washed 3× with KPL wash buffer (1× diluted in deionized water) (SeraCare, Milford, MA, USA) and blocked overnight at 4 °C with 1% BSA/KPL buffer. Lung homogenates treated with PMSF Protease Inhibitor (Thermo Fisher Scientific, Grand Island, NY, USA), standards, and controls were added to the plates and incubated overnight at 4 °C. 12 h later the wells were washed 3× with KPL wash buffer and a biotinylated detection antibody was added for 2 h at room temperature (RT). Wells were washed and incubated with streptavidin-HRP in the dark for 20 min at RT. Wells were washed 3× with KPL wash buffer and detected with One-Step TMB (ThermoFisher) and stopped with Stop Solution (ThermoFisher). Plates were read on a BioTek plate reader at OD₄₅₀. A standard curve was used to quantify protein concentrations using standards included in commercial kits.

Bergeron H.C., Murray J., Arora A., Nuñez Castrejon A.M., DuBois R.M., Anderson L.J., Kauvar L.M, & Tripp R.A. (2023). Immune Prophylaxis Targeting the Respiratory Syncytial Virus (RSV) G Protein. Viruses, 15(5), 1067.

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Quantification of IFNβ and IFNλ2/λ3 in Biological Samples

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IFNβ and IFNλ2/λ3 capture ELISAs were performed according to the manufacturer’s instructions (R&D Systems, Minneapolis, MN). Briefly, the capture antibody was coated onto a high-binding ELISA plate (Corning) overnight. The following day, wells were washed 3x with KPL wash buffer [1X diluted in deionized water (SeraCare)] and blocked overnight at 4°C with 1% BSA/KPL buffer. BALF and cell supernatants were added to the plates and incubated overnight at 4°C. Twelve hours later, the wells were washed 3x with KPL wash buffer, and a biotinylated detection antibody was added for 2 h at room temperature. Wells were washed and incubated with streptavidin-HRP in the dark for 20 min at room temperature. Wells were washed 3x with KPL wash buffer and detected with One-Step TMB (ThermoFisher) and stopped with Stop Solution (ThermoFisher). Plates were read on a BioTek plate reader at OD₄₅₀. A standard curve was generated to quantify protein concentrations using standards included in respective kits.

Bergeron H.C., Kauvar L.M, & Tripp R.A. (2023). Anti-G protein antibodies targeting the RSV G protein CX3C chemokine region improve the interferon response. Therapeutic Advances in Infectious Disease, 10, 20499361231161157.

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