Xbridge beh130

Manufactured by Waters Corporation

The XBridge BEH130 is a high-performance liquid chromatography (HPLC) column designed for analytical separations. It features a silica-based stationary phase with a particle size of 1.7 μm and a pore size of 130 Å, providing high-resolution separations of a wide range of analytes.

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Lab products found in correlation

Dionex ultimate 3000 hplc system, by Thermo Fisher Scientific (2 mentions) 1525 binary pump, by Waters Corporation (1 mentions) Mercury 300 mhz, by Agilent Technologies (1 mentions)

Close spelling variants of this product

XBridge BEH130 Prep C18 XBridge BEH130 column XBridge BEH130 C18 column BEH130

3 protocols using xbridge beh130

High pH Reversed-Phase Fractionation

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For high pH reversed phase fractionation, dried samples were reconstituted in MS-grade water with 10% fractionation buffer A (25 mM ammonium bicarbonate, pH 8) and centrifuged for 5 min at 20,000x g and 4°C. The supernatant was then loaded on a C18 column (XBridge BEH130, 3.5µm, 2.1×150mm, Waters Corp), which was connected to a Dionex Ultimate 3000 HPLC system (Thermo Fisher). After injecting 200 µg protein digest of the sample at a flow rate of 200 µg/min, the system was equilibrated for 5 min with 85% fractionation buffer B (MS-grade water), 10% fractionation buffer A and 5% fractionation buffer C (ACN). Peptides were eluted in a three-step linear gradient from 5% to 9% buffer C in 2 min with a constant amount of 10% buffer A. Then, a linear gradient from 9% to 47% buffer C in 91 min and from 47% to 55% buffer C in 3 min (with buffer A being constant at 10%) was used.
Starting from minute 3, 96 fractions (1 fraction/min) were collected in a 96-well plate and pooled to 48 fractions. For that, column 7 was pooled to column 1, column 8 was pooled to column 2 and so forth. All fractions were frozen at −80°C and dried using a Speed-Vac.

Philippi A., Heller S., Costa I.G., Senée V., Breunig M., Li Z., Kwon G., Russell R., Illing A., Lin Q., Hohwieler M., Degavre A., Zalloua P., Liebau S., Schuster M., Krumm J., Zhang X., Geusz R., Benthuysen J.R., Wang A., Chiou J., Gaulton K., Neubauer H., Simon E., Klein T., Wagner M., Nair G., Besse C., Dandine-Roulland C., Olaso R., Deleuze J.F., Kuster B., Hebrok M., Seufferlein T., Sander M., Boehm B.O., Oswald F., Nicolino M., Julier C, & Kleger A. (2021). ONECUT1 mutations and variants in diabetes. Nature medicine, 27(11), 1928-1940.

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Synthetic Peptide Preparation and Purification

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All reagents and solvents were acquired from Sigma-Aldrich (Milano, Italy). Solution phase peptide synthesis was applied to prepare the final products KA1, K2–K6 as TFA salts, following the procedures reported below. Boc-protected intermediates were purified by silica gel column chromatography where necessary, or by trituration in Et₂O. Final products KA1, K2–K6 were purified by RP–HPLC on a Waters XBridge BEH130 (C18 5.0 μm, 250 × 10 mm column; flow rate of 7 mL/min; Waters Binary pump 1525; eluent: linear gradient of H₂O/ACN 0.1% TFA, ranging from 5% to 95% ACN in 32 min). The purity of the N^α-Boc-protected products was confirmed by NMR analysis on a Varian Mercury 300 MHz. The purity of all final compounds was assessed by NMR analysis, ESI–LRMS, and by analytical RP–HPLC (C18-bonded 4.6 × 150 mm; flow rate of 1 mL/min; eluent: gradient of H₂O/ACN 0.1% TFA, ranging from 5% to 95% ACN in 26 min, recorded at 254, 275, and 213 nm and was found to be ≥95%). The mass spectrometry (MS) equipment was composed as follows: LCQ Thermo Finnigan ion trap mass spectrometer (San Jose, CA, USA) with an electrospray ionization (ESI) source; capillary temperature: 300 °C; spray voltage: 4.00 kV; nitrogen (N₂) as the sheath and auxiliary gas.

Szűcs E., Stefanucci A., Dimmito M.P., Zádor F., Pieretti S., Zengin G., Vécsei L., Benyhe S., Nalli M, & Mollica A. (2020). Discovery of Kynurenines Containing Oligopeptides as Potent Opioid Receptor Agonists. Biomolecules, 10(2), 284.

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High pH Reversed Phase Fractionation

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For high pH reversed phase fractionation, dried samples were reconstituted in MS-grade water with 10% fractionation buffer A (25 mM ammonium bicarbonate (pH 8)) and centrifuged for 5 min at 20.000 × g and 4°C. The supernatant was then loaded on a C18 column (XBridge BEH130, 3.5 μm, 2.1×150 mm, Waters Corp), which was connected to a Dionex Ultimate 3000 HPLC system (Thermo). After injecting 100 μg peptides at a flow rate of 200 μg/min, the system was equilibrated for 5 min with 85% fractionation buffer B (MS-grade water), 10% fractionation buffer A and 5% fractionation buffer C (ACN). Peptides were eluted in a three-step linear gradient from 5% to 7% buffer C in 1 min with a constant amount of 10% buffer A. Then, a linear gradient from 7% to 42% buffer C in 44 min and from 42% to 80% buffer C in 6 min (with buffer A being constant at 10%) was used.
Starting from minute 3, 48 fractions (1 fraction/min) were collected in a 96-well plate and pooled to 24 fractions. For that, column 4 was pooled to column 1, column 5 was pooled to column 2 and column 6 was pooled to column 3. All fractions were frozen at −80°C and dried using a Speed-Vac.

Breunig M., Merkle J., Wagner M., Melzer M.K., Barth T.F., Engleitner T., Krumm J., Wiedenmann S., Cohrs C.M., Perkhofer L., Jain G., Krüger J., Hermann P.C., Schmid M., Madácsy T., Varga Á., Griger J., Azoitei N., Müller M., Wessely O., Robey P.G., Heller S., Dantes Z., Reichert M., Günes C., Bolenz C., Kuhn F., Maléth J., Speier S., Liebau S., Sipos B., Kuster B., Seufferlein T., Rad R., Meier M., Hohwieler M, & Kleger A. (2021). Modelling Plasticity and Dysplasia of Pancreatic Ductal Organoids Derived from Human Pluripotent Stem Cells. Cell stem cell, 28(6), 1105-1124.e19.

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