Minidawn treos detector

Manufactured by Wyatt Technology

Sourced in United States

The MiniDAWN TREOS detector is a light scattering instrument designed to measure the molar mass and size of macromolecules in solution. It features a three-angle static light scattering detector that provides accurate and reliable data for a wide range of sample types.

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45 protocols using minidawn treos detector

SEC-MALS Analysis of Protein Samples

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SEC-MALS measurements were carried out with a miniDAWN TREOS detector (Wyatt Technology Corporation) coupled to an Agilent 1260 Infinity HPLC. A total of 240–360 µg protein samples were injected into a size exclusion chromatography column (ENrich SEC 650, Bio-Rad) and continuously run at a flow rate of 0.5 ml min⁻¹ in the buffer containing 20 mM Tris-HCl, pH 7.5, 200 mM NaCl, 2 mM DTT, and 0.02% NaN₃. The molecular weight was determined by multi-angle laser light scattering using an in-line miniDAWN TREOS detector and an Optilab T-rEX differential refractive index detector (Wyatt Technology Corporation). Bovine serum albumin (Sigma, A1900) was used for system calibration and the data were analyzed using ASTRA 6 Software (Wyatt Technology Corporation) with the dn/dc value set to 0.185 ml g⁻¹. Experiment was repeated two times with similar observations, and a representative figure is shown.

Su M.Y., Som N., Wu C.Y., Su S.C., Kuo Y.T., Ke L.C., Ho M.R., Tzeng S.R., Teng C.H., Mengin-Lecreulx D., Reddy M, & Chang C.I. (2017). Structural basis of adaptor-mediated protein degradation by the tail-specific PDZ-protease Prc. Nature Communications, 8, 1516.

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Protein Purification by Size Exclusion Chromatography

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The KlNmd4 protein (100 µL at 1 mg/mL) was injected at a flow rate of 0.75 mL/min on a Superdex TM 200 Increase 10/300 GL column (GE-Healthcare) using buffer B. Elution was followed by a UV-visible spectrophotometer, a MiniDawn TREOS detector (Wyatt Technology) and a RID-20A refractive index detector (Shimadzu). Data were processed with the program ASTRA 6.1 (Wyatt Technology). The M w was directly calculated from the absolute light scattering measurements using a dn/dc value of 0.183.

Barbarin-Bocahu I., & Graille M. (2021). Artificial intelligence to solve the X-ray crystallography phase problem: a case study report.

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Molecular Weight Determination by SEC

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The weight-average molecular weights (Mw) of each sample were determined through Size Exclusion Chromatography (SEC) measurements using two columns in series from Shodex SB 806 M HQ associated with a pre-column SB-G 6B. The solvent used was 0.1 mol L -1 NaNO3 including 0.3 g L -1 NaN3. A laser light scattering Mini Dawn Treos detector from Wyatt was associated with a Wyatt differential refractometer. The selected flow rate was 0.5 mL min -1 at 30 °C, the dn/dc was equal to 0.155 and sample concentration injected was 4 mg mL -1 . The samples were dissolved in water under stirring during 24 h and filtrated on a 0.2 m porous membrane before injection.

Amaral S.d.C., Roux D., Marlu R., Rinaudo M., Barbieri S.F., & Silveira J.L.M. (2021). Extraction, characterization and gelling ability of pectins from Araçá (Psidium cattleianum Sabine) fruits. Food Hydrocolloids, 121, 106845-106845.

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RIG-I Stem dsRNA Interaction Characterization

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E373A RIG-I (300 nM) was incubated with stem dsRNA (100 nM) for 30 min on ice. The complex was injected onto Superdex 200 Increase (GE Healthcare) Size Exclusion Chromatography (SEC) in the buffer of 50mM HEPES pH 7.5, 50mM NaCl, 1 mM MgCl₂, 5mM DTT and 500μM ATP with a flow rate of 0.5 mL/min. The complex was characterized by SEC coupled with Multi-Angle Light Scattering (SEC-MALS). Sample eluted from the SEC passed through a miniDAWN TREOS detector (Wyatt Technology) and an Optilab T-rEX refractometer (Wyatt Technology). Light scattering intensity and eluent refractive index were recorded and analyzed by ASTRA v7.0.1.24 software to yield a weight-averaged molecular mass.

Devarkar S.C., Schweibenz B., Wang C., Marcotrigiano J, & Patel S.S. (2018). RIG-I uses an ATPase-powered translocation-throttling mechanism for kinetic proofreading of RNAs and oligomerization. Molecular cell, 72(2), 355-368.e4.

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Molecular Mass and Oligomeric State Analysis

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The molecular mass and the oligomeric state of PETISCO, its subunits and other proteins in solution were determined by size exclusion chromatography (SEC) coupled to multi-angle light scattering (MALS). Individual proteins or protein complexes were analyzed at concentrations between 2 and 5 mg/mL in a buffer consisting of 20 mM Tris/HCl pH 7.5, 150 mM NaCl, 2 mM DTT. A Superdex 200 Increase 10/300 GL column (GE Healthcare Life Sciences) was connected to a 1260 Infinity HPLC system (Agilent Technologies) coupled to a MiniDawn Treos detector (Wyatt Technologies) with a laser emitting at 690 nm. An RI-101 detector (Shodex) was used for refractive index measurement. Data analysis was performed using Astra 7 software package (Wyatt Technologies).

Perez-Borrajero C., Podvalnaya N., Holleis K., Lichtenberger R., Karaulanov E., Simon B., Basquin J., Hennig J., Ketting R.F., & Falk S. (2021). Structural basis of PETISCO complex assembly during piRNA biogenesis inC. elegans.

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Determining Protein Oligomeric States by SEC-MALS

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The molecular mass and the oligomeric state of PETISCO, its subunits, and other proteins in solution were determined by size exclusion chromatography (SEC) coupled to multiangle light scattering (MALS). Individual proteins or protein complexes were analyzed at concentrations between 2 and 5 mg/mL in a buffer consisting of 20 mM Tris/HCl (pH 7.5), 150 mM NaCl, 2 mM DTT. A Superdex 200 Increase 10/300 GL column (GE Healthcare Life Sciences) was connected to a 1260 Infinity HPLC system (Agilent Technologies) coupled to a MiniDawn Treos detector (Wyatt Technologies) with a laser emitting at 690 nm. An RI-101 detector (Shodex) was used for refractive index measurement. Data analysis was performed using the Astra 7 software package (Wyatt Technologies).

Perez-Borrajero C., Podvalnaya N., Holleis K., Lichtenberger R., Karaulanov E., Simon B., Basquin J., Hennig J., Ketting R.F, & Falk S. (2021). Structural basis of PETISCO complex assembly during piRNA biogenesis in C. elegans. Genes & Development, 35(17-18), 1304-1323.

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Size-exclusion chromatography and MALS analysis

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Size-exclusion chromatography experiments coupled to multi-angle laser light scattering (MALS) and refractometry (RI) were performed on a Superdex S200 Increase 5/150 GL column with size-exclusion buffer 50 mM Tris pH 8, 150 mM NaCl for DARPin K5 and CagI:K5 complex and with buffer 50 mM Tris pH 8, 200 mM NaCl, 5% glycerol v/v for others CagI:DARPin complexes. Fifty microliters of proteins were injected at a concentration of 5 to 8 mg/mL. Online MALS detection was performed with a miniDAWN-TREOS detector (Wyatt Technology Corp., Santa Barbara, CA, USA) using a laser emitting at 690 nm and by refractive index measurement using an Optilab T-rEX system (Wyatt Technology Corp.). Weight-averaged molar masses (Mw) were calculated using the ASTRA software (Wyatt Technology Corp.).

Blanc M., Lettl C., Guérin J., Vieille A., Furler S., Briand-Schumacher S., Dreier B., Bergé C., Plückthun A., Vadon-Le Goff S., Fronzes R., Rousselle P., Fischer W, & Terradot L. (2023). Designed Ankyrin Repeat Proteins provide insights into the structure and function of CagI and are potent inhibitors of CagA translocation by the Helicobacter pylori type IV secretion system. PLOS Pathogens, 19(5), e1011368.

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Size-Exclusion Chromatography of Sgk3 Protein

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Sgk3 (80 μl) at 0.9 mg/ml was injected onto Superdex 200 Increase 10/300 GL column (Cytiva) equilibrated in 20 mM Tris pH 8, 150 mM NaCl, 1% (v/v) glycerol, 1 mM TCEP. Separation was carried out using HPLC Agilent Technologies 1260 infinity with a flow rate of 0.5 ml/min operated at room temperature. Online light scattering of 690 nm laser was recorded on a miniDawn Treos detector (Wyatt Technology Corp.). Shodex RI-101 (Shodex) refractive index detector was used to measure protein concentration online. Data was analyzed using the ASTRA software (Wyatt Technology Corp.).

Pokorny D., Truebestein L., Fleming K.D., Burke J.E, & Leonard T.A. (2021). In vitro reconstitution of Sgk3 activation by phosphatidylinositol 3-phosphate. The Journal of Biological Chemistry, 297(2), 100919.

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SEC-MALS Analysis of Purified B3GNT2

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Purified B3GNT2 (20 μl at 1 mg/ml) was analyzed by SEC-MALS on a Superdex 75 gel filtration column (GE Life Sciences) in a buffer containing 25 mM HEPES (pH 7.4), 150 mM NaCl, 0.02% NaN₃. In-line light scattering was measured using a MiniDAWN TREOS detector (Wyatt Technology) and differential refractive index using an Optilab rEX detector (Wyatt Technology). Data were analyzed using Astra 6 software (Wyatt Technology).

Kadirvelraj R., Yang J.Y., Kim H.W., Sanders J.H., Moremen K.W, & Wood Z.A. (2020). Comparison of human poly-N-acetyl-lactosamine synthase structure with GT-A fold glycosyltransferases supports a modular assembly of catalytic subsites. The Journal of Biological Chemistry, 296, 100110.

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Size Exclusion Chromatography of EXT1-2

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The EXT1–2 preparation (1 mg ml⁻¹) was injected on an analytical scale Superdex 75 gel filtration column in a buffer containing 20 mM HEPES (pH 7.4) and 150 mM NaCl. In-line light scattering was measured using a MiniDAWN TREOS detector (Wyatt Technology) and differential refractive index using a Optilab rEx detector (Wyatt Technology). Data analysis was performed using the ASTRA software package 6.0 (Wyatt Technology).

Watts C.R, & Vanryckeghem M. (2001). Laryngeal dysfunction in Amyotrophic Lateral Sclerosis: a review and case report. BMC Ear, Nose, and Throat Disorders, 1, 1.

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