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Easy nlc 1200 uhplc system

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The EASY-nLC 1200 UHPLC system is a high-performance liquid chromatography instrument designed for analytical separations. The system features a modular design, precise solvent delivery, and high-pressure capabilities, enabling the separation and analysis of complex samples. The EASY-nLC 1200 is suitable for a variety of applications, including proteomics, metabolomics, and small molecule analysis.

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Lab products found in correlation

Q exactive hf x mass spectrometer, by Thermo Fisher Scientific (15 mentions) Reprosil pur c18 aq beads, by Dr. Maisch (13 mentions) Picofrit 10 mm opening, by New Objective (6 mentions) Orbitrap q exactive hf x mass spectrometer, by Thermo Fisher Scientific (6 mentions) Orbitrap fusion lumos, by Thermo Fisher Scientific (6 mentions) Irt peptide, by Biognosys (5 mentions) Thermo fusion lumos mass spectrometer, by Thermo Fisher Scientific (5 mentions) Thermo fusion lumos, by Thermo Fisher Scientific (4 mentions) Picofrit 10 μm, by New Objective (3 mentions) Trypsin, by Promega (3 mentions) 2 μm c18 beads, by Thermo Fisher Scientific (3 mentions) Orbitrap fusion lumos mass spectrometer, by Thermo Fisher Scientific (3 mentions) Picofrit 10 μm opening, by New Objective (2 mentions) Reprosil pur 120 c18 aq resin, by Dr. Maisch (2 mentions) Proteome discoverer, by Thermo Fisher Scientific (2 mentions) Nanospray flex ion, by Thermo Fisher Scientific (2 mentions) Q exactive hybrid quadrupole orbitrap mass spectrometer, by Thermo Fisher Scientific (2 mentions) Hcd fragmentation cell, by Thermo Fisher Scientific (2 mentions) Nanou3000uhplc system, by Thermo Fisher Scientific (2 mentions) Q exactive mass spectrometer, by Thermo Fisher Scientific (2 mentions)

Spelling variants of this product

EASY-nLC 1200 (450 citations) Easy nLC (237 citations) EASY-nLC system (111 citations) UHPLC system (58 citations) EASY-nLCTM 1200 UHPLC system (43 citations)

55 protocols using easy nlc 1200 uhplc system

1

DIA-based Quantitative Proteomics of ccRCC Samples

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Unlabeled, digested peptide material from individual tissue samples (ccRCC and NAT) was spiked with index Retention Time (iRT) peptides (Biognosys) and subjected to data-independent acquisition (DIA) analysis. Peptides (~0.8 μg) were separated on an Easy nLC 1200 UHPLC system (Thermo Scientific) on an in-house packed 20 cm x 75 μm diameter C18 column (1.9 μm Reprosil-Pur C18-AQ beads (Dr. Maisch GmbH); Picofrit 10 μm opening (New Objective)). The column was heated to 50°C using a column heater (Phoenix-ST). The flow rate was 0.200 μl/min with 0.1% formic acid and 3% acetonitrile in water (A) and 0.1% formic acid, 90% acetonitrile (B). The peptides were separated with a 7%–30% B gradient in 84 mins and analyzed using the Thermo Fusion Lumos mass spectrometer (Thermo Scientific). The DIA segment consisted of one MS1 scan (350-1650 m/z range, 120K resolution) followed by 30 MS2 scans (variable m/z range, 30K resolution). Additional parameters were as follows: MS1: RF Lens – 30%, AGC Target 4.0e5, Max IT – 50 ms, charge state include - 2-6; MS2: isolation width (m/z) – 0.7, AGC Target - 2.0e5, Max IT – 120 ms.
Clark D.J., Dhanasekaran S.M., Petralia F., Pan J., Song X., Hu Y., da Veiga Leprevost F., Reva B., Lih T.S., Chang H.Y., Ma W., Huang C., Ricketts C.J., Chen L., Krek A., Li Y., Rykunov D., Li Q.K., Chen L.S., Ozbek U., Vasaikar S., Wu Y., Yoo S., Chowdhury S., Wyczalkowski M.A., Ji J., Schnaubelt M., Kong A., Sethuraman S., Avtonomov D.M., Ao M., Colaprico A., Cao S., Cho K.C., Kalayci S., Ma S., Liu W., Ruggles K., Calinawan A., Gümüş Z.H., Geiszler D., Kawaler E., Teo G.C., Wen B., Zhang Y., Keegan S., Li K., Chen F., Edwards N., Pierorazio P.M., Chen X.S., Pavlovich C.P., Hakimi A.A., Brominski G., Hsieh J.J., Antczak A., Omelchenko T., Lubinski J., Wiznerowicz M., Linehan W.M., Kinsinger C.R., Thiagarajan M., Boja E.S., Mesri M., Hiltke T., Robles A.I., Rodriguez H., Qian J., Fenyö D., Zhang B., Ding L., Schadt E., Chinnaiyan A.M., Zhang Z., Omenn G.S., Cieslik M., Chan D.W., Nesvizhskii A.I., Wang P, & Zhang H. (2019). Integrated Proteogenomic Characterization of Clear Cell Renal Cell Carcinoma. Cell, 179(4), 964-983.e31.
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2

Comprehensive Proteomic Profiling Workflow

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Global proteome and phosphoproteome fractions were analyzed using the same instrumentation and methodology. Peptides (~0.8 μg) were separated on an Easy nLC 1200 UHPLC system (Thermo Scientific) on an in-house packed 20 cm x 75 mm diameter C18 column (1.9 mm Reprosil-Pur C18-AQ beads (Dr. Maisch GmbH); Picofrit 10 mm opening (New Objective)). The column was heated to 50°C using a column heater (Phoenix-ST). The flow rate was 0.200 μl/min with 0.1% formic acid and 2% acetonitrile in water (A) and 0.1% formic acid, 90% acetonitrile (B). The peptides were separated with a 6%–30% B gradient in 84 min and analyzed using the Thermo Fusion Lumos mass spectrometer (Thermo Scientific). Parameters were as follows: MS1: resolution – 60,000, mass range – 350 to 1800 m/z, RF Lens – 30%, AGC Target 4.0e5, Max IT – 50 ms, charge state include - 2-6, dynamic exclusion – 45 s, top 20 ions selected for MS2; MS2: resolution-50,000, high-energy collision dissociation activation energy (HCD)-37, isolation width (m/z) – 0.7, AGC Target – 2.0e5, Max IT – 105 ms.
Clark D.J., Dhanasekaran S.M., Petralia F., Pan J., Song X., Hu Y., da Veiga Leprevost F., Reva B., Lih T.S., Chang H.Y., Ma W., Huang C., Ricketts C.J., Chen L., Krek A., Li Y., Rykunov D., Li Q.K., Chen L.S., Ozbek U., Vasaikar S., Wu Y., Yoo S., Chowdhury S., Wyczalkowski M.A., Ji J., Schnaubelt M., Kong A., Sethuraman S., Avtonomov D.M., Ao M., Colaprico A., Cao S., Cho K.C., Kalayci S., Ma S., Liu W., Ruggles K., Calinawan A., Gümüş Z.H., Geiszler D., Kawaler E., Teo G.C., Wen B., Zhang Y., Keegan S., Li K., Chen F., Edwards N., Pierorazio P.M., Chen X.S., Pavlovich C.P., Hakimi A.A., Brominski G., Hsieh J.J., Antczak A., Omelchenko T., Lubinski J., Wiznerowicz M., Linehan W.M., Kinsinger C.R., Thiagarajan M., Boja E.S., Mesri M., Hiltke T., Robles A.I., Rodriguez H., Qian J., Fenyö D., Zhang B., Ding L., Schadt E., Chinnaiyan A.M., Zhang Z., Omenn G.S., Cieslik M., Chan D.W., Nesvizhskii A.I., Wang P, & Zhang H. (2019). Integrated Proteogenomic Characterization of Clear Cell Renal Cell Carcinoma. Cell, 179(4), 964-983.e31.
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3

Proteomics Analysis by DIA-MS

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The proteomics analysis was conducted with an Orbitrap QE HF mass spectrometer coupled to an EASY-nLC 1200 UHPLC system (Thermo Fisher Scientific, Waltham, MA, USA). The peptides were loaded onto a trap column and separated on an analytical column (75 µm × 500 mm, Kyoto Monotech, Kyoto, Japan). Peptides were eluted using a 60 min gradient from 5 to 30% buffer B (0.1% FA in 99.9% acetonitrile) at a flow rate of 300 nl/min. Moreover, all samples were spiked with iRT peptides for alignment of retention times across the samples. The mass spectrometer was operated in DIA mode, and a full-scan MS spectrum (350–1500 m/z) was collected with a resolution of 120,000. The injection time was less than 100 ms, and the automatic gain control (AGC) target value was 3e6.
Chen Z., Wu J., Wang W., Tang X., Zhou L., Lv Y, & Zheng Y. (2023). Investigation of the Pathogenic Mechanism of Ciprofloxacin in Aortic Aneurysm and Dissection by an Integrated Proteomics and Network Pharmacology Strategy. Journal of Clinical Medicine, 12(4), 1270.
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4

Targeted Proteomic Analysis of Proteins

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For proteins that were not detected in TMT‐based proteomic analysis, parallel reaction monitoring (PRM)‐based proteomic analysis was used to specifically search and analyse the proteins. Briefly, the proteins were extracted and enzymatically hydrolysed similar to the above method. Same amount of trypsin treated‐peptide was taken of each sample, and labelled peptide GAGSSEPVTGLDAK as an internal standard of each sample and the peptides were dissolved in solvent A (0.1% formic acid in 2% acetonitrile)and solvent B (0.1% formic acid in 80% acetonitrile). PRM mass spectrometric analysis was performed using an EASY‐nLC™ 1200 UHPLC system (Thermo Fisher Scientific, USA). The full scan mass spectrum resolution was set to 60 000 (200 m/z), the maximum C‐trap and IT were 3 × 106 and 50 ms, respectively. The PRM resolution was set to 30 000 (200 m/z), the maximum C‐trap and IT were 1 × 106 and 100 ms, respectively. Normalized collision energy is 28. The data were analysed using the Skyline software (MacCoss Lab, Univ. of Washington), and the peak area was corrected using the internal standard peptide.
Feng L., Zhang W., Shen Q., Miao C., Chen L., Li Y., Gu X., Fan M., Ma Y., Wang H., Liu X, & Zhang X. (2021). Bile acid metabolism dysregulation associates with cancer cachexia: roles of liver and gut microbiome. Journal of Cachexia, Sarcopenia and Muscle, 12(6), 1553-1569.
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5

Orbitrap-based Quantitative Proteomics and Glycoproteomics

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For data acquisition, peptide samples were analyzed using an Orbitrap Lumos Fusion system. Peptide (1 μg) was separated using an EASY-nLC 1200 UHPLC system (Thermo Scientific) on an in-house packed 20 cm × 75 μm diameter C18 column (1.9 μm ReproSil-Pur C18-AQ beads (Dr. Maisch GmbH); Picofrit 10 μm opening (New Objective)). The column was heated to 50 °C using a column heater (Phoenix-ST). The flow rate was 0.300 μL/min with 0.1% formic acid and 2% acetonitrile in water (A) and 0.1% formic acid and 90% acetonitrile (B). The peptides were separated with a 6–30% B gradient in 84 min and analyzed using a Thermo Fusion Lumos mass spectrometer (Thermo Scientific). Parameters were as follows: MS1: resolution, 60,000; mass range, 350–800 m/z, RF lens, 30%, AGC target, 4.0 × 105; Max IT, 50 ms; charge state, 2–6; dynamic exclusion, 45 s; top 20 ions selected for MS2; MS2: resolution, 50,000; high-energy collision dissociation activation energy (HCD), 34 (global peptide) or 36 (intact glycopeptide); isolation width (m/z), 0.7; AGC target, 2.0 × 105; Max IT, 105 ms. Three replicates were analyzed for the proteomic and glycoproteomic analyses.
Clark D.J., Schnaubelt M., Hoti N., Hu Y., Zhou Y., Gooya M, & Zhang H. (2020). Impact of Increased FUT8 Expression on the Extracellular Vesicle Proteome in Prostate Cancer Cells.. Journal of proteome research, 19(6), 2195-2205.
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6

Proteomic Profiling using EASY-nLC™ 1200

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Proteomic analyses were performed using an EASY-nLC™ 1200 UHPLC system (ThermoFisher Scientific, Germany) coupled to an Orbitrap Q Exactive HF-X mass spectrometer (ThermoFisher) operating in the data-dependent acquisition (DDA) mode which was carried out as previously described (Pang et al., 2020 (link)).
Zeng F., Pang H., Chen Y., Zheng H., Li W., Ramanathan S., Hoare R., Monaghan S.J., Lin X, & Jian J. (2021). First Succinylome Profiling of Vibrio alginolyticus Reveals Key Role of Lysine Succinylation in Cellular Metabolism and Virulence. Frontiers in Cellular and Infection Microbiology, 10, 626574.
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7

Peptide Separation and Mass Spectrometry

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One µg of peptide was separated using Easy nLC 1200 UHPLC system (Thermo Scientific) on an in-house packed 20 cm × 75 μm diameter C18 column (1.9 μm Reprosil-Pur C18-AQ beads (Dr. Maisch GmbH); Picofrit 10 μm opening (New Objective)). The column was heated to 50 °C using a column heater (Phoenix-ST). The flow rate was 0.200 μl/min with 0.1% formic acid and 2% acetonitrile in water (A) and 0.1% formic acid, 90% acetonitrile (B). The peptides were separated with a 6-30% B gradient in 84 min and analyzed using the Thermo Fusion Lumos mass spectrometer (Thermo Scientific). Parameters were as followed MS1: resolution – 60,000, mass range – 350 to 1800 m/z, RF Lens – 30%, AGC Target 4.0e5, Max IT – 50 ms, charge state include - 2-6, dynamic exclusion – 45 s, top 20 ions selected for MS2; MS2: resolution – 15,000, high-energy collision dissociation activation energy (HCD) – 34 (nonTMT-labeled samples) or 37 (TMT-labeled samples), isolation width (m/z) – 0.7, AGC Target – 5.0e4.
Theodros D., Murter B.M., Sidhom J.W., Nirschl T.R., Clark D.J., Chen L., Tam A.J., Blosser R.L., Schwen Z.R., Johnson M.H., Pierorazio P.M., Zhang H., Ganguly S., Pardoll D.M, & Zarif J.C. (2020). High-dimensional Cytometry (ExCYT) and Mass Spectrometry of Myeloid Infiltrate in Clinically Localized Clear Cell Renal Cell Carcinoma Identifies Novel Potential Myeloid Targets for Immunotherapy. Molecular & Cellular Proteomics : MCP, 19(11), 1850-1859.
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8

Proteomic Analysis of Digested HDL Proteins

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Digested HDL proteins (50 ng) were loaded onto a trap column (nanoViper C18, 3 μm, 75 μm × 2 cm, Thermo Scientific) and eluted onto a C18 column (nanoViper, 2 μm, 75 μm × 15 cm, Thermo Scientific). Peptides were analyzed using an Easy-nLC 1200 UHPLC system (Thermo Scientific) coupled to an Orbitrap Fusion Lumos (Thermo Scientific) equipped with a nanospray FlexNG ion source (Thermo Scientific) in a 44 min gradient and normalized collision energy of 30 for HCD fragmentation. For untargeted analysis (DDA), peptides were analyzed using MS1 resolution of 120,000 (at m/z 200) with AGC target set to 4 × 105, m/z range of 350–1550, and maximum injection time of 50 ms. MS2 resolution was set at 30,000 (at m/z 200) with AGC target of 5 × 104 and maximum injection time of 54 ms. For targeted analysis (DIA), peptides were quantified using Orbitrap resolution of 30,000 (at m/z 200) with AGC target of 5 × 105, precursor m/z range of 400–900, scan range of product ions between m/z 100 and 1000, maximum injection time of 54 ms, and isolation windows of 25 m/z with 0.5 m/z margins.
Holzer M., Ljubojevic-Holzer S., Souza Junior D.R., Stadler J.T., Rani A., Scharnagl H., Ronsein G.E, & Marsche G. (2022). HDL Isolated by Immunoaffinity, Ultracentrifugation, or Precipitation is Compositionally and Functionally Distinct. Journal of Lipid Research, 63(12), 100307.
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9

Shotgun Proteomics with Orbitrap Q Exactive HF-X

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An EASY-nLC™ 1200 UHPLC system (Thermo Fisher Scientific, Germany) coupled to an Orbitrap Q Exactive HF-X mass spectrometer (Thermo Fisher Scientific, Germany) was used to perform Shotgun proteomics analyses in the data-dependent acquisition (DDA) mode. The specific details have been described previously (Pang et al., 2019 (link)).
Wang J., Pang H., Yin L., Zeng F., Wang N., Hoare R., Monaghan S.J., Li W, & Jian J. (2022). A Comprehensive Analysis of the Lysine Acetylome in the Aquatic Animals Pathogenic Bacterium Vibrio mimicus. Frontiers in Microbiology, 13, 816968.
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10

Quantitative Proteomic Analysis of Neuronal Secretome

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All mass spectrometry measurements were conducted on an EASY-nLC 1200 UHPLC system (Thermo Fisher Scientific) connected to a Q-Exactive™ HF Hybrid Quadrupole-Orbitrap™ mass spectrometer (Thermo Fisher Scientific) as described (39 (link)). Peptides were separated on self-packed 30 cm long C18 columns (ReproSil-Pur 120 C18-AQ resin Dr. Maisch GmbH) with a diameter of 75 μm. CSF samples were analyzed using data-dependent acquisition (DDA) using the MaxQuant software (40 (link)) (1% FDR on protein and peptide level). Whereas hiSPECS secretome samples were analyzed using data-independent acquisition (DIA) in combination with self-generated hiSPECS neuronal secretome library (35 (link)) using Spectronaut Pulsar (41 (link)). Details of the chromatography (120-min gradient) and mass spectrometry parameters were set as described (35 (link)). Proteomic data are accessible via PRIDE with the following project accession: PXD028096.
Tüshaus J., Müller S.A., Shrouder J., Arends M., Simons M., Plesnila N., Blobel C.P, & Lichtenthaler S.F. (2021). The pseudoprotease iRhom1 controls ectodomain shedding of membrane proteins in the nervous system. FASEB journal : official publication of the Federation of American Societies for Experimental Biology, 35(11), e21962.
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