Prior to analysis, sample gel pieces from three different transformants expressing the Flag-UPF2 allele were washed, reduced, alkylated, and in-gel tryptic digested. Additional analysis was performed by in-gel gluc-c digestion. Proteolytic peptides were extracted from the gel. The phosphopeptides were enriched by TiO2 TopTip (Glygen Corp., Columbia, MD, USA). Both flow-through and elute were analyzed by LC-MS/MS, respectively. For this analysis, an ABSCIEX QSTAR XL mass spectrometer (AB Sciex, Framingham, MA, USA) was used. This MS was coupled to a CapLC (Waters Corp., Milford, MA, USA) HPLC with a Phenomenex C18 (3 μm, 300 A, 100 μm ID × 150 mm, Phenomenex, Torrence, CA, USA) analytical column, and a Vydac Everest C18, 300 A, 5 μm trap column. The solvents used were composed of 5% CH3CN + 0.1% formic acid + 0.01% TFA (Solvent A) and 85% CH3CN + 10% isopropanol + 5% H2O + 0.1% formic acid + 0.01% TFA (Solvent B). Samples were kept at a 250nl/min flow rate and a 60 min linear gradient from 10% to 100% B.
Absciex qstar xl
Manufactured by AB Sciex
Sourced in United States
The ABSCIEX QSTAR XL is a high-performance quadrupole time-of-flight (QTOF) mass spectrometer. It is designed to provide accurate and reliable mass analysis for a wide range of applications in the life sciences and analytical chemistry fields.
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Lab products found in correlation
2 protocols using absciex qstar xl
1
Phosphoproteome Analysis of UPF2 Allele
2
Characterization of Novel Compounds
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Unless otherwise mentioned,
commercially available reagents were purchased from Sigma-Aldrich
and Acros Organics without further purification. Solvent purification
was according to Purification of Laboratory Chemicals.700 All reactions were carried out under
Ar atmosphere and were monitored by thin layer chromatography (TLC)
using precoated Merck silica gel 60 F254 alumina plates (0.25 mm).
Visualization was accomplished using ultraviolet light (256 and 365
nm). Column chromatography was carried out using silica gel (230–400
mesh) supplied by Merck. 1H, 13C, 19F, 31P, and 11B NMR spectra were recorded at
298 K on a Varian Unity 400 spectrometer; the chemical shifts (δ
in ppm) are reported with DMSO-d6 (δ
2.50 for 1H NMR and δ 39.52 for 13C NMR),
CDCl3 (δ 7.26 for 1H NMR and δ 77.06
for 13C NMR), or CD3OD (δ 3.31 for 1H NMR and δ 49.00 for 13C NMR) as internal
references. Single crystal X-ray diffraction data was collected on
a Bruker D8 VENTURE diffractometer equipped with Oxford Cryostream
800+. High-resolution mass spectra (HRMS) were obtained using the
electrospray ionization (ESI) method or fast atom bombardment (FAB)
method with AB SCIEX QSTAR XL.
commercially available reagents were purchased from Sigma-Aldrich
and Acros Organics without further purification. Solvent purification
was according to Purification of Laboratory Chemicals.700 All reactions were carried out under
Ar atmosphere and were monitored by thin layer chromatography (TLC)
using precoated Merck silica gel 60 F254 alumina plates (0.25 mm).
Visualization was accomplished using ultraviolet light (256 and 365
nm). Column chromatography was carried out using silica gel (230–400
mesh) supplied by Merck. 1H, 13C, 19F, 31P, and 11B NMR spectra were recorded at
298 K on a Varian Unity 400 spectrometer; the chemical shifts (δ
in ppm) are reported with DMSO-d6 (δ
2.50 for 1H NMR and δ 39.52 for 13C NMR),
CDCl3 (δ 7.26 for 1H NMR and δ 77.06
for 13C NMR), or CD3OD (δ 3.31 for 1H NMR and δ 49.00 for 13C NMR) as internal
references. Single crystal X-ray diffraction data was collected on
a Bruker D8 VENTURE diffractometer equipped with Oxford Cryostream
800+. High-resolution mass spectra (HRMS) were obtained using the
electrospray ionization (ESI) method or fast atom bombardment (FAB)
method with AB SCIEX QSTAR XL.
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