Acquity m class nanolc system

Manufactured by Waters Corporation

Sourced in United States

The Acquity M-class nanoLC system is a high-performance liquid chromatography instrument designed for the separation and analysis of small-volume samples. It features a modular design, adjustable flow rates, and precise temperature control to ensure reliable and reproducible results.

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Lab products found in correlation

Picofrit column, by New Objective (4 mentions) Magic c18aq resin, by Bruker (3 mentions) Q exactive plus mass spectrometer, by Thermo Fisher Scientific (3 mentions) Nanoease symmetry c18 trapping column, by Waters Corporation (2 mentions) Nanoease symmetry c18 trapping column, by New Objective (2 mentions) Synapt g2si instrument, by Waters Corporation (2 mentions) Orbitrap analyzer, by Thermo Fisher Scientific (1 mentions) Unifi scientific information system, by Waters Corporation (1 mentions) Masslynx 4, by Waters Corporation (1 mentions) Q exactive plus, by Thermo Fisher Scientific (1 mentions)

6 protocols using acquity m class nanolc system

Nanoflow LC-MS/MS Proteomics Pipeline

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An Acquity M-class nanoLC system (Waters, Milford, MA, USA) was used, loading 5 µL of the sample (1 mg) at a rate of 15 mL/min for 3 min onto a nanoEase Symmetry C18 trapping column (180 mm × 20 mm). It was then washed onto a PicoFrit column (75 mm ID × 250 mm; New Objective, Woburn, MA, USA) packed with Magic C18AQ resin (Michrom Bioresources, Auburn, CA, USA). The column was then eluted of peptides into the Q Exactive Plus mass spectrometer (Thermofisher Scientific, NSW, Australia) using the following program: 5%–30% MS buffer B (98% acetonitrile + 0.2% formic acid) for 90 min, 30%–80% MS buffer B for 3 min, 80% MS buffer B for 2 min, 80%–5% for 3 min. The peptides that were eluted were ionised at 2000 V. A data-dependent MS/MS (dd-MS2) experiment was performed with a 350–1500 Da survey scan performed at a resolution of 70,000 m/z for peptides of charge state 2+ or higher with an automatic gain control (AGC) target of 3 × 106 and a 50 ms maximum injection time. The top 12 peptides were selectively fragmented in the higher-energy collisional dissociation (HCD) cell using a 1.4 m/z isolation window, an AGC target of 1 × 105 and a 100 ms maximum injection time. The fragments were scanned in the Orbitrap analyser at a resolution of 17,500 and the product ion fragment masses were measured over a 120–2000 Da mass range. The mass of the precursor peptide was then excluded for 30 s.

Fajardo J., Milthorpe B.K, & Santos J. (2020). Molecular Mechanisms Involved in Neural Substructure Development during Phosphodiesterase Inhibitor Treatment of Mesenchymal Stem Cells. International Journal of Molecular Sciences, 21(14), 4867.

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Nanoflow LC-MS/MS Proteomics Workflow

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Using an Acquity M-class nanoLC system (Waters, Milford, MA, USA), 5 —L of the sample (1 —g) was loaded at 15 —L/min for 3 min onto a nanoEase Symmetry C18 trapping column (180 —m x 20 mm) before being washed onto a PicoFrit column (75 —m ID x 250 mm; New Objective, Woburn, MA, USA) packed with Magic C18AQ resin (Michrom Bioresources, Auburn, CA, USA). Peptides were eluted from the column and into the source of a Q Exactive™ Plus mass spectrometer (Thermo Scientific, Rockford, IL, USA) using the following program: 5–30% MS buffer B (98% acetonitrile + 0.2% formic acid) over 90 min, 30–80% MS buffer B over 3 min, 80% MS buffer B for 2 min, 80–5% for 3 min. The eluting peptides were ionized at 2000V. A Data Dependent MS/MS (dd-MS²) experiment was performed, with a survey scan of 350–1500 Da performed at 70,000 resolution for peptides of charge state 2+ or higher with an AGC target of 3e6 and maximum Injection Time of 50 ms. The Top 12 peptides were selectively fragmented in the HCD cell using an isolation window of 1.4 m/z, an AGC target of 1e5 and maximum injection time of 100 ms. Fragments were scanned in the Orbitrap analyzer (Thermo Scientific, Rockford, IL, USA) at 17,500 resolution and the product ion fragment masses measured over a mass range of 120–2000 Da. The mass of the precursor peptide was then excluded for 30 s.

Roediger B., Lee Q., Tikoo S., Cobbin J.C., Henderson J.M., Jormakka M., O’Rourke M.B., Padula M.P., Pinello N., Henry M., Wynne M., Santagostino S.F., Brayton C.F., Rasmussen L., Lisowski L., Tay S.S., Harris D.C., Bertram J.F., Dowling J.P., Bertolino P., Lai J.H., Wu W., Bachovchin W.W., Wong J.J., Gorrell M.D., Shaban B., Holmes E.C., Jolly C.J., Monette S, & Weninger W. (2018). An atypical parvovirus driving chronic tubulointerstitial nephropathy and kidney fibrosis. Cell, 175(2), 530-543.e24.

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High-Resolution Mass Spectrometry for Peptide Analysis

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The high-resolution mass spectrometry analysis was performed using a Synapt-G2Si instrument coupled to the Acquity M-class nano-LC system (Waters, Prague, Czech Republic). The HSS T3 column (100 Å, 1.8 µm, 75µm × 250 mm) was used for peptide separation. The column was heated to 40°C. Mobile phase A was composed of 0.1% formic acid in deionized water (v/v), and mobile phase B consisted of acetonitrile with 0.1% formic acid (v/v). The mobile phase gradient program was as follows: 2% B in 0–5 min, 2–40% B in 5–35 min and then 2% B in 35–50 min. The flow rate was 0.3 µL min⁻¹, and the injection volume was 1 µL (final concentration 100 fmol). Data were acquired with MassLynx 4.2 software and analyzed by UNIFI Scientific Information System (Waters, Prague, Czech Republic).

Majerova P., Hanes J., Olesova D., Sinsky J., Pilipcinec E, & Kovac A. (2020). Novel Blood–Brain Barrier Shuttle Peptides Discovered through the Phage Display Method. Molecules, 25(4), 874.

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Nanoflow LC-MS/MS Peptide Profiling

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Using an Acquity M-class nanoLC system (Waters, Milford, MA, USA), 5 µL of the sample was loaded at 15 μL/min for 3 min onto a nanoEase Symmetry C18 trapping column (180 μm × 20 mm) before being washed onto a PicoFrit column (75 μm ID × 350 mm; New Objective, Woburn, MA, USA) packed with SP-120-1.7-ODS-BIO resin (1.7 μm, Osaka Soda Co., Tokyo, Japan) heated to 45 °C at 300 nL/min. Peptides were eluted from the column and into the source of a Q Exactive Plus mass spectrometer (Thermo Scientific, Carlsbad, CA, USA) using the following program: 5–30% MS buffer B (98% acetonitrile + 0.2% formic acid) over 90 min, 30–80% MS buffer B over 3 min, 80% MS buffer B for 2 min, 80–5% for 3 min. The eluting peptides were ionised at 2400 V. A data-dependant MS/MS (dd-MS2) experiment was performed, with a survey scan of 350–1500 Da performed at 70,000 resolution for peptides of charge state 2+ or higher with an AGC target of 3 × 10⁶ and maximum injection time of 50 ms. The top 12 peptides were selected fragmented in the HCD cell using an isolation window of 1.4 m/z, an AGC target of 1 × 10⁵ and maximum injection time of 100 ms. Fragments were scanned in the Orbitrap analyser at 17,500 resolution, and the product ion fragment were masses measured over a mass range of 120–2000 Da. The mass of the precursor peptide was then excluded for 30 s.

Gomila Pelegri N., Stanczak A.M., Bottomley A.L., Milthorpe B.K., Gorrie C.A., Padula M.P, & Santos J. (2023). Adipose-Derived Stem Cells Spontaneously Express Neural Markers When Grown in a PEG-Based 3D Matrix. International Journal of Molecular Sciences, 24(15), 12139.

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Proteomic Analysis Using Nanoflow LC-MS/MS

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An Acquity M-class nanoLC system (Waters, USA) was used, loading 5 µL of the sample (1 mg) at a rate of 15 mL/min for 3 min onto a nanoEase Symmetry C18 trapping column (180 mm × 20 mm). It was then washed onto a PicoFrit column (75 mm ID × 250 mm; New Objective, Woburn, MA, USA) packed with Magic C18AQ resin (Michrom Bioresources, Auburn, CA, USA). The column was then eluted of peptides into the Q Exactive Plus mass spectrometer (Thermofisher Scientific, NSW, Australia) using the following program: 5%–30% MS buffer B (98% Acetonitrile +0.2% Formic Acid) over 90 min, 30%–80% MS buffer B over 3 min, 80% MS buffer B for 2 min, 80%–5% for 3 min. The peptides that were eluted were ionised at 2000 V. A data dependant MS/MS (dd-MS2) experiment was performed, with a 350–1500 Da survey scan was performed at a resolution of 70,000 m/z for peptides of charge state 2+ or higher with an Automatic Gain Control (AGC) target of 3 × 10⁶ and a 50 ms maximum injection time. The top 12 peptides were selectively fragmented in the Higher-energy collisional dissociation (HCD) cell using a 1.4 m/z isolation window, an AGC target of 1 × 105 and a 100 ms maximum injection time. The fragments were scanned in the Orbitrap analyser at a resolution of 17,500 and the product ion fragment masses were measured over a 120–2000 Da mass range. The mass of the precursor peptide was then excluded for 30 s.

Santos J., Hubert T, & Milthorpe B.K. (2020). Valproic Acid Promotes Early Neural Differentiation in Adult Mesenchymal Stem Cells Through Protein Signalling Pathways. Cells, 9(3), 619.

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High-resolution nanoLC-MS/MS analysis

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The high-resolution mass spectrometry analysis was performed using a Synapt-G2Si instrument coupled to an Acquity M-class nano-LC system and equipped with an ionKey source (Waters Corporation, Milford, MA, USA). A separation device (chip) iKey Peptide BEH (100Å, 18 µm, 150 µm × 100 mm) was used for separation. The iKey device was heated to 40 °C. Mobile phase A was composed of formic acid in MPW (0.1%, v/v), and mobile phase B consisted of acetonitrile with 0.1% formic acid (v/v). A mobile phase gradient program was as follows: 10% B (0–1 min), increasing to 90% B (1–15 min) and then returning to 10% B and re-equilibrating at 17.1 to 20 min. The flow rate was 3 µL·min⁻¹ and the injection volume was 3 µL. In a mass spectrometer, nitrogen was used as a drying gas and high purity nitrogen (N2) was used as a collision gas for collision-induced dissociation (CID). The mass spectrometric parameters were as follows: positive ion mode; desolvation gas flow 800 L·h⁻¹; desolvation temperature 350 °C; capillary voltage 2.8 kV; source temperature 100 °C; mass range, m/z 50–1200 for MS1.

Galba J., Piešťanský J., Kováč A., Olešová D., Cehlár O., Kertys M., Kozlík P., Chaľová P., Tirčová B., Slíž K, & Mikuš P. (2021). Fast and Sensitive Screening of Oxandrolone and Its Major Metabolite 17-Epi-Oxandrolone in Human Urine by UHPLC—MS/MS with On-Line SPE Sample Pretreatment. Molecules, 26(2), 480.

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