Reprosil pur c18 aq 3 μm

Manufactured by Dr. Maisch

Sourced in Germany

Reprosil-Pur C18-AQ (3 μm) is a reversed-phase high-performance liquid chromatography (HPLC) column. It features a 3 μm particle size and a C18-AQ stationary phase, which is designed for the separation of a wide range of polar and non-polar analytes.

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Close spelling variants of this product

ReproSil-Pur C18-AQ (72 citations) ReproSil-Pur C18-AQ 3 μm resin (22 citations) Reprosil-Pur C18 (21 citations) RP ReproSil-Pur C18-AQ 3 μm resin (4 citations) Reprosil-AQ Pur (3 citations) Reprosil-Pur 120 C18-AQ 3 μm reversed-phase material (3 citations) ReproSil-Pur 130 C18-AQ 3 μm particles (2 citations)

7 protocols using reprosil pur c18 aq 3 μm

Profiling Cytosine and Methylcytosine by nLC-MS

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nLC was configured with a two-column system consisting of a 300 μm ID x 0.5 cm C18 trap column (Dionex) and a 75 μm ID x 25 cm Reprosil-Pur C18-AQ (3 μm; Dr. Maisch GmbH, Germany) analytical nano-column packed in-house using a Dionex RSLC Ultimate 3000 (Thermo Scientific, San Jose, CA, USA). nLC was coupled online to an Orbitrap Fusion Lumos mass spectrometer (Thermo Scientific). The spray voltage was set to 2.3 kV and the temperature of the heated capillary was set to 275 °C. The full scan range was 110–600 m/z acquired in the Orbitrap at a resolution 120,000. The source fragmentation energy was set at 30 V and RF lens % was set at 50. Extracted signals are the protonated nucleobases Cytosine (C) and methylcytosine (mC) with m/z at 112.0505 and 126.0662, respectively.

Sun Y., Stransky S., Aguilan J., Koul S., Garforth S.J., Brenowitz M, & Sidoli S. (2021). High Throughput and Low Bias DNA Methylation and Hydroxymethylation Analysis by Direct Injection Mass Spectrometry. Analytica chimica acta, 1180, 338880.

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Peptide Nano-LC-MS/MS Workflow

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Peptides were loaded directly (2%
B, 500 nL/min) onto a spray emitter analytical column (75 μm
inner diameter, 8 μm opening, 250 mm length; New Objectives)
packed with C18 material (ReproSil-Pur C18-AQ 3 μm; Dr Maisch
GmbH, Ammerbuch-Entringen, Germany) using an air pressure pump (Proxeon
Biosystems).^{37 (link)} The 0.1% formic acid served
as mobile phase A and 0.1% formic acid/80% acetonitrile as mobile
phase B. Peptides were eluted (200 nL/min, linear gradient of 2–40%
B over 139 min) directly into an Orbitrap Fusion Lumos Tribrid mass
spectrometer (Thermo Fisher Scientific, San Jose, CA). Survey spectra
were recorded in the Orbitrap at 120000 resolution. Spectra for all
fragmentation methods were acquired using a scan range of 300–1700 m/z. Precursor ion isolation was performed
with the quadrupole and an m/z window
of 1.6 Th. The precursor automatic gain control (AGC) target value
was 4 × 10⁵, maximum injection time 50 ms. For CID
only, CID collision energy was set to 30%. For HCD only, HCD collision
energy was set to 35%. For ETD only, the option to inject ions for
all available parallelizable time was selected (anion AGC 5 ×
10⁴, 60 ms maximum injection time). Supplemental activation
(SA) collision energy was set to 10% for ETciD, and 25% for EThcD.

Giese S.H., Belsom A, & Rappsilber J. (2016). Optimized Fragmentation Regime for Diazirine Photo-Cross-Linked Peptides. Analytical Chemistry, 88(16), 8239-8247.

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Hybrid Mass Spectrometry for Peptide Analysis

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SCX-Stage-Tip fractions were analyzed using a hybrid linear ion trap-Orbitrap mass spectrometer (LTQ-Orbitrap Velos, Thermo Fisher Scientific, Bremen Germany) applying a “high-high” acquisition strategy. Peptides were separated on an analytical column that was packed with C18 material (ReproSil-Pur C18-AQ 3 μm; Dr. Maisch GmbH, Ammerbuch-Entringen, Germany) in a spray emitter (75-μm inner diameter, 8-μm opening, 250-mm length; New Objectives, Woburn, MA) (39 (link)). Mobile phase A consisted of water and 0.5% v/v acetic acid. Mobile phase B consisted of acetonitrile and 0.5% v/v acetic acid. Peptides were loaded at a flow-rate of 0.6 μl/min and eluted at 0.3 μl/min using a linear gradient going from 3% mobile phase B to 35% mobile phase B over 130 min, followed by a linear increase from 35% to 80% mobile phase B in 5 mins. The eluted peptides were directly introduced into the mass spectrometer. MS data were acquired in the data-dependent mode. For each acquisition cycle, the mass spectrum was recorded in the Orbitrap with a resolution of 100,000. The eight most intense ions with a precursor charge state 3+ or greater were fragmented in the linear ion trap by collision-induced disassociation (CID). The fragmentation spectra were then recorded in the Orbitrap at a resolution of 7,500. Dynamic exclusion was enabled with single repeat count and 60-s exclusion duration.

Chen Z.A., Pellarin R., Fischer L., Sali A., Nilges M., Barlow P.N, & Rappsilber J. (2016). Structure of Complement C3(H2O) Revealed By Quantitative Cross-Linking/Mass Spectrometry And Modeling. Molecular & Cellular Proteomics : MCP, 15(8), 2730-2743.

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Nano-HPLC-MS Analysis of SCX Fractions

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Each SCX fraction was analyzed via online
C18-nano-HPLC-MS with a system consisting of an Easy-nLC 1000 gradient
HPLC (Thermo Scientific) and a Q-Exactive mass spectrometer (Thermo
Scientific). Fractions were injected into a homemade precolumn (100
μm × 15 mm; Reprosil-Pur C18-AQ 3 μm, Dr. Maisch),
equilibrated with solvent A (100/0.1 water/formic acid (FA) v/v),
and eluted using a homemade analytical nano-HPLC column (15 cm ×
50 μm; Reprosil-Pur C18-AQ 3 um). The gradient was run from
0 to 30% solvent B (100/0.1 acetonitrile/formic acid (FA) v/v) in
120 min. The nano-HPLC column was drawn to a ∼5 μm tip
and acted as the electrospray needle of the MS source. The Q-Exactive
mass spectrometer was operated in top10-mode. Parameters were as follows:
a resolution of 70 000 at an AGC target value of 3 million
with a maximum fill time of 20 ms (full scan) and a resolution of
17 500 at an AGC target value of 100 000 with a maximum
fill time of 60 ms for MS/MS (normalized collision energy (NCE) 27%,
dynamic exclusion 30 s) at an intensity threshold of 17 000.
The apex trigger was set to 1–5 s and allowed charges were
2–5.

Uribe M.L., Martín-Nieto J., Quereda C., Rubio-Fernández M., Cruces J., Janssen G.M., de Ru A.H., van Veelen P.A, & Hensbergen P.J. (2021). Retinal Proteomics of a Mouse Model of Dystroglycanopathies Reveals Molecular Alterations in Photoreceptors. Journal of Proteome Research, 20(6), 3268-3277.

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iTRAQ-Based Proteomics Analysis Pipeline

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The iTRAQ labelled sample was applied onto an EASY nano-LC system (Proxeon Biosystems, Odense, Denmark). The peptides were loaded directly onto a 20-cm 100-μm inner diameter, 360-μm outer diameter, ReproSil – Pur C18 AQ 3-μm (Dr. Maisch, Germany) homemade reversed phase capillary column. The peptides were eluted from the column into an LTQ-orbitrap XL mass spectrometer (Thermo Fisher Scientific, Bremen, Germany), using a gradient from 100% phase A (0.1 % formic acid) to 34 % phase B (0.1 % formic acid, 90%acetonitrile) in 120 min. at 250 nl min⁻¹. The data-dependent analysis was performed using one MS full scan in the area 400 Da – 1800 Da performed in the Orbitrap with 30000 in resolution, followed by CID and HCD of the three most intense ions (Rewitz et al., 2009 (link)) The threshold for ion-selection was 10000 and the conditions for CID fragmentation were normalized collision energy 35, isolation width 2.0, activation time 30 ms, and for HCD fragmentation normalized collision energy 50, isolation width 2.0, activation time 1 ms.

Frederiksen S.B., Holm L.L., Larsen M.R., Doktor T.K., Andersen H.S., Hastings M.L., Hua Y., Krainer A.R, & Andresen B.S. (2020). Identification of SRSF10 as a regulator of SMN2 ISS-N1. Human mutation, 42(3), 246-260.

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Nano-LC-MS/MS Proteomic Analysis Protocol

Cited 1 time

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Samples were analyzed by using a nanoLC-MS/MS setup. NanoLC was configured with a 75 μm ID x 15 cm Reprosil-Pur C18-AQ (3 μm; Dr. Maisch GmbH, Germany) nano-column using an EASY-nLC nano-HPLC (Thermo Scientific, San Jose, CA, USA), packed in-house. The HPLC gradient was as follows: 2% to 28% solvent B (A = 0.1% formic acid; B = 95% MeCN, 0.1% formic acid) over 45 minutes, from 28% to 80% solvent B in 5 minutes, 80% B for 10 minutes at a flow-rate of 300 nL/min. nanoLC was coupled to an LTQ Velos mass spectrometer (Thermo Scientific, San Jose, CA, USA). For DIA, two full scan MS spectra (m/z 300−1100) were acquired in the ion trap within a DIA duty cycle, and 16 ms/ms were performed with an isolation window of 50 Da. Normalized collision energy (CE) was set to 35% with activation Q of 0.25^{12 (link)}.

Guo Q., Sidoli S., Garcia B.A, & Zhao X. (2017). Assessment of quantification precision of histone post-translational modifications by using an ion trap and down to 50,000 cells as starting material. Journal of proteome research, 17(1), 234-242.

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Phosphoproteome Analysis via nanoLC-MS/MS

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Phosphopeptide-enriched samples were analyzed via on-line C18-nano-HPLC-MS with a system consisting of an Easy nLC 1000 gradient HPLC system (Thermo, Bremen, Germany), and a Q-Exactive mass spectrometer (Thermo). Fractions were injected onto a homemade precolumn (100 μm × 15 mm; Reprosil-Pur C18-AQ 3 μm, Dr. Maisch, Ammerbuch, Germany) and eluted via a homemade analytical nano-HPLC column (15 cm × 50 μm; Reprosil-Pur C18-AQ 3 um). The gradient was run from 0% to 30% solvent B (10/90/0.1 (v/v/v) water/ACN/FA) in 120 min. The nano-HPLC column was drawn to a tip of ∼5 μm and acted as the electrospray needle of the MS source. The Q-Exactive mass spectrometer was operated in top10-mode. Parameters were resolution 70,500 at an AGC target value of 3,000,000, maximum fill time of 250 ms (full scan), and resolution 17,500 at an AGC target value of 200,000/maximum fill time of 80 ms for MS/MS at an intensity threshold of 2,500. Apex trigger was set to 1 to 15 seconds, and allowed charges were 2–6. Each sample was analyzed in duplo.

Treffers E.E., Tas A., Scholte F.E., de Ru A.H., Snijder E.J., van Veelen P.A, & van Hemert M.J. (2023). The alphavirus nonstructural protein 2 NTPase induces a host translational shut-off through phosphorylation of eEF2 via cAMP-PKA-eEF2K signaling. PLOS Pathogens, 19(2), e1011179.

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