Nanoacquity uplc system

Manufactured by Waters Corporation

Sourced in United States, United Kingdom, Germany

The NanoAcquity UPLC system is a high-performance liquid chromatography (HPLC) instrument designed for ultra-high performance liquid chromatography (UPLC) applications. The system is capable of separating and analyzing complex mixtures with high resolution and sensitivity.

Automatically generated - may contain errors

Lab products found in correlation

333 protocols using nanoacquity uplc system

Label-free Proteomic Analysis Pipeline

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Label-free proteomic analysis was performed in a nanoACQUITY UPLC system (Waters, Milford, MA) coupled to a Xevo Q-TOF G2 mass spectrometer (Waters, Milford, MA), as described elsewhere (Lobo, Leite et al. 2015). The nanoACQUITY UPLC system is equipped with a Trap Columm (100Å, 5 µm, 180 µm × 200 mm) and a HSS T3 M-Class type column (analytical column 75 μm × 150 mm; 1.8 μm) (Waters, Milford, MA). ProteinLynx GlobalServer software (PLGS) version 3.03 (Waters, Milford, MA) was used to process and search the LC-MSE continuum data.
Peptides identification and difference in expression among the groups was obtained using the Protein Lynx Global Server (PLGS) software (version 3.03, Waters Co., UK) as described elsewhere (Lima Leite, Gualiume Vaz Madureira Lobo et al. 2014 (link)). The procedures and bioinformatics analysis were performed as described previously (Dionizio, Melo et al. 2018, Dionizio, Melo et al. 2020 ).

Dionizio A., Uyghurturk D.A., Souza Melo C.G., Sabino-Arias I.T., Araujo T.T., Ventura T.M., Martins Perles J.V., Zanoni J.N., Den Besten P, & Rabelo Buzalaf M.A. (2021). Intestinal changes associated with fluoride exposure in rats: Integrative morphological, proteomic and microbiome analyses. Chemosphere, 273, 129607.

+ Open protocol

+ Expand

Proteomic Analysis of Biological Samples

Cited 2 times

Check if the same lab product or an alternative is used in the 5 most similar protocols

The label-free proteomic analysis was performed in a nanoACQUITY UPLC system (Waters, Milford, MA, USA) coupled to a Xevo Q-TOF G2 mass spectrometer (Waters, Milford, MA, USA). The nanoACQUITY UPLC system was equipped with a Trap Columm (100 Å, 5 μm, 180 μm × 200 mm) and a HSS T3 M-Class type column (analytical column 75 μm × 150 mm; 1.8 μm) (Waters, Milford, MA, USA). The reading and identification of peptides was performed using the ProteinLynx GlobalServer software (PLGS) version 3.03 (Waters, Milford, MA, USA), as previously described [63 (link)]. The PLGS software, applying the Monte-Carlo algorithm, was used to determine the difference in protein expression between the groups, considering p < 0.05 for downregulated proteins and 1 − p > 0.95 for upregulated proteins. The identification of proteins was performed by downloading UniProt databases. Then, bioinformatics analyses were performed using Cytoscape^® 3.6 (Java^®) with the ClusterMarker plugin for the PPI network, and for the determination of the biological process groups based on Gene Ontology annotations, we used the ClueGO plugin [64 (link)].

Souza-Monteiro D., Guerra M.C., Bittencourt L.O., Aragão W.A., Dionizio A., Silveira F.M., Buzalaf M.A., Martins M.D., Crespo-Lopez M.E, & Lima R.R. (2022). Salivary Glands after Prolonged Aluminum Exposure: Proteomic Approach Underlying Biochemical and Morphological Impairments in Rats. International Journal of Molecular Sciences, 23(4), 2251.

+ Open protocol

+ Expand

Nanoflow UPLC-MS/MS Proteomic Analysis

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Tryptic
peptides were analyzed with a nanoAcquity UPLC system coupled to a
Synapt G2-Si HDMS mass spectrometer with a nanoelectrospray ionization
source (Waters Corporation, Manchester, UK). The nanoAcquity UPLC
system consisted of a C18, 5 μm, 180 μm × 20 mm trap
column and an HSS-T3 C18 1.8 μm, 75 μm × 250 mm analytical
column (Waters Corporation, Manchester, UK) set to trapping mode.
Samples containing 300 ng of protein were injected in each run. Mobile
phases A and B consisted of 0.1% formic acid and 3% dimethyl sulfoxide
in water (v/v) and 0.1% formic acid and 3% dimethyl sulfoxide in acetonitrile
(v/v), respectively. Peptide separation was done using a gradient
from 3% to 40% (v/v) of mobile phase B at a constant flow rate of
0.3 μL/min over 120 min. Lock-mass correction was performed
by spraying a lock-mass solution containing [Glu1]-fibrinopeptide
B (0.1 μM) and leu-enkephalin (1 μM) through the reference
channel every 60 s. Data were acquired using a data-independent acquisition
workflow in positive ion mode with a UDMS^E method.^{19 (link)−21 (link)} The system’s performance and stability were monitored by
injecting a commercially available HeLa digest (Thermo Scientific,
Waltham, MA) after every seventh sample injection.

Sandbaumhüter F.A., Nezhyva M., Andrén P.E, & Jansson E.T. (2023). Label-Free Quantitative Thermal Proteome Profiling Reveals Target Transcription Factors with Activities Modulated by MC3R Signaling. Analytical Chemistry, 95(41), 15400-15408.

+ Open protocol

+ Expand

Quantifying Tau Phosphorylation in CSF

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

CSF was acquired to evaluate phosphorylation of tau at pT181, pS202, pT205, and pT217 using corresponding (p-tau/t-tau) ratios. Further details on CSF preparation has been previously described (Barthelemy et al., 2020a (link)). Tau phosphorylation was analyzed by nano liquid chromatography-high-resolution mass spectrometry (nanoLC-MS/HRMS) using Parallel Reaction Monitoring with Higher-energy collisional dissociation (HCD) fragmentation. NanoLC-MS/HRMS experiments were performed using a nanoAcquity UPLC system (Waters, Mildford, Massachusetts) coupled to a Fusion Tribrid mass spectrometer (Thermo Scientific, San Jose, California). CSF Tau phosphorylation ratios were calculated using measured ratios between MS/HRMS transitions of endogenous unphosphorylated peptides and 15 N labeled peptides from protein internal standard. Each phosphorylated/unphosphorylated peptide endogenous ratio was normalized using the ratio measured on the MS/HRMS transitions of the corresponding AQUA phosphorylated/unphosphorylated peptide internal standards (Barthelemy et al., 2020a (link)).

Strain J.F., Barthelemy N., Horie K., Gordon B.A., Kilgore C., Aschenbrenner A., Cruchaga C., Xiong C., Joseph-Mathurin N., Hassenstab J., Fagan A.M., Li Y., Karch C.M., Perrin R.J., Berman S.B., Chhatwal J.P., Graff-Radford N.R., Mori H., Levin J., Noble J.M., Allegri R., Schofield P.R., Marcus D.S., Holtzman D.M., Morris J.C., Benzinger T.L., McDade E.M., Bateman R.J, & Ances B.M. (2022). CSF Tau phosphorylation at Thr205 is associated with loss of white matter integrity in autosomal dominant Alzheimer disease. Neurobiology of disease, 168, 105714.

+ Open protocol

+ Expand

Quantitative Proteomics by LC-MS/MS

Check if the same lab product or an alternative is used in the 5 most similar protocols

LC-MS/MS data were acquired on a Q-Exactive mass spectrometer (Thermo Scientific) coupled with a nanoAcquity UPLC system (Waters). Seven μL of peptide samples were injected and separated by a custom packed analytical C18 column (70 cm × 75 μm i.d., 3 um particle size of Jupiter C18, Phenomenex) in a 3 h gradient. The flow rate of the mobile phase (buffer A: 0.1% formic acid in water; buffer B: 0.1% formic acid in acetonitrile) was 300 nL/min. For peptide elution buffer B was increased to 12% at 36 min, 30% at 135 min, 45% at 175 min and finally 95% at 180 min. A data-dependent acquisition method was applied, in which a full MS scan was followed by up to 12 data-dependent MS/MS scans of the most abundant peptide precursors. A dynamic exclusion of 30 s was implemented to prevent repeated sequencing of previously selected peptides. For MS scans (400–2000 m/z; resolution of 35000 at 400 m/z), the automatic gain control (AGC) was set to 3-e⁶ with the maximum injection time (IT) of 20 ms. An isolation window of 2 m/z in the quadrupole was used for selecting precursor ions, which were fragmented by higher-energy collisional dissociation (HCD) at a normalized collision energy of 35. For MS/MS scans (200–2000 m/z with first fixed mass of 100 m/z; resolution of 17500 at 400 m/z), the AGC was set to 1-e⁵ and maximum IT was 100 ms.

Cao X., Zhang T., DeLoid G.M., Gaffrey M.J., Weitz K.K., Thrall B.D., Qian W.J, & Demokritou P. (2020). Evaluation of the cytotoxic and cellular proteome impacts of food-grade TiO2 (E171) using simulated gastrointestinal digestions and a tri-culture small intestinal epithelial model. NanoImpact, 17, 10.1016/j.impact.2019.100202.

+ Open protocol

+ Expand

LC-MS Analysis of ShHTL7 Enzymatic Reactions

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

For LC-MS analysis: a 200 μL reaction solution (100 mM HEPES, 150 mM NaCl, pH 7.0) containing 10, or 400 μM of tested samples 4a, (R)- and (S)-4a, and 4g-i, and 100 μg of recombinant ShHTL7 or its mutants was incubated at 25 °C for 30 min. Subsequently, the solution was filtered and analyzed by a nanoACQUITY UPLC system, which was directly interfaced with an SYNAPT-G2-Si mass spectrometer produced by Waters Company. For LC-MS/MS analysis, the above chemicals treated ShHTL7 reaction solution was subjected to the SDS-PAGE analysis. The corresponding protein band around 30 KDa was excised, respectively, then digested with trypsin (Promega) in NH₄HCO₃ solution (50 mM) at 37 °C for 24 h^{23 (link),26 (link)–28 (link)}. The covalent modification of C₅H₅O₂/C₆H₆O on the peptide was analyzed by a Thermo-Dionex Ultimate 3000 HPLC system, which was directly interfaced with a Thermo Orbitrap Fusion Lumos mass spectrometer. MS-MS spectra are generated by pLabel software^{53 (link),54 (link)}.

Wang D., Pang Z., Yu H., Thiombiano B., Walmsley A., Yu S., Zhang Y., Wei T., Liang L., Wang J., Wen X., Bouwmeester H.J., Yao R, & Xi Z. (2022). Probing strigolactone perception mechanisms with rationally designed small-molecule agonists stimulating germination of root parasitic weeds. Nature Communications, 13, 3987.

+ Open protocol

+ Expand

Phosphoproteome Profiling by LC-MS/MS

Check if the same lab product or an alternative is used in the 5 most similar protocols

Sample preparation of the liquid chromatography-tandem mass spectrometry (LC MS/MS) analysis was performed following the previously described protocol [44 (link)]. Briefly, immunoprecipitated proteins were subjected to Dithiothreitol (DTT) reduction, Iodoacetamide (IAN)-mediated alkylation followed by trypsin digestion. The digested sample was desalted by Spin Desalting column (Thermo) and acidified with 0.5% Trifluoroacetic acid (TFA), 50% acetonitrile then subjected to titanium dioxide enrichment using the Top Tips system (Glygen Corp). The resulting phosphopeptide-enriched sample, dissolved in 70% formic acid and diluted with 0.1% TFA, was then subjected to LC-MS/MS analysis using the Orbitrap Fusion Mass Spectrometer that is equipped with a Waters nanoACQUITY UPLC system. A Waters Symmetry C18 180 μm x 20 mm trap column and a 1.7 μm, 75 μm x 250 mm nanoACQUITY UPLC column was utilized for online peptide separation. The acquired data was peak picked and searched using the Mascot Distiller and the Mascot search algorithm, respectively. Manual examination of the MS/MS spectra (as shown in Fig 1B) and the corresponding assigned fragment ions were conducted to verify the identified phosphopeptide.

Li J., Walsh A., Lam T.T., Delecluse H.J, & El-Guindy A. (2019). A single phosphoacceptor residue in BGLF3 is essential for transcription of Epstein-Barr virus late genes. PLoS Pathogens, 15(8), e1007980.

+ Open protocol

+ Expand

Mass Spectrometry-based Protein Identification

Check if the same lab product or an alternative is used in the 5 most similar protocols

Selected protein spots were excised from the gels and distained, reduced with 10 mM dithiothreitol in 25 mM ammonium bicarbonate followed by protein alkylation with 55 mM iodoacetamide. Protein digestion was carried out overnight at 37°C with sequencing grade trypsin (Promega, Madison, WI, USA). Nanoscale LC separation of tryptic peptides was performed with a nanoACQUITY UPLC System (Waters, Milford, MA, USA) and tandem mass spectrometry analysis carried out in a SYNAPT HDMS (Waters) as previously reported (Huerta-Ocampo et al., 2014 (link)) with a brief modification: Accurate mass data were collected in an alternating data dependent acquisition mode (DDA). In low energy mode, data were collected at constant collision energy of 3 eV. In elevated-energy mode, the collision energy was ramped from 15 to 45 eV during 5 s of integration.

Dufoo-Hurtado M.D., Huerta-Ocampo J.Á., Barrera-Pacheco A., Barba de la Rosa A.P, & Mercado-Silva E.M. (2015). Low temperature conditioning of garlic (Allium sativum L.) “seed” cloves induces alterations in sprouts proteome. Frontiers in Plant Science, 6, 332.

+ Open protocol

+ Expand

HPLC-MS/MS Analysis of PhIP

Check if the same lab product or an alternative is used in the 5 most similar protocols

The analysis was performed with a nanoAcquity UPLC system (Waters Corporation) equipped with a BEH Shield RP18 column (0.3 × 150 mm, 1.7 μm, Waters Corporation) and a HESI II source interfaced with a TSQ Quantiva^™ TQ mass spectrometer (Thermo Scientific, San Jose, CA). Solvent A was 5 mM NH₄HCO₃ (pH 9.0) and solvent B was CH₃CN. A linear gradient was employed, starting at 10% B and arriving to 99% B in 10 min at a flow rate 5 μL/min.. The MS instrument parameters were: positive spray voltage, 3000 V; Nitrogen was the sheath gas, 4 arbitrary units; no auxiliary gas and sweep gas; ion transfer tube temperature, 400 °C; dwell time, 10 ms; Q1 and Q3 resolution (FWHM), 0.7; Argon was the CID gas, 1.5 mTorr; source fragmentation, 10 V. The selected reaction monitoring (SRM) scan transitions were: PhIP, 225.1 > 210.1, 140.1; and [²H₃C]-PhIP, 228.1 > 210.1, 140.1.

Guo J., Yonemori K., Le Marchand L, & Turesky R.J. (2015). A Method to Biomonitor the Cooked Meat Carcinogen 2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) in Dyed Hair by Ultra Performance Liquid Chromatography-Orbitrap High Resolution Multistage Mass Spectrometry. Analytical chemistry, 87(12), 5872-5877.

+ Open protocol

+ Expand

Shotgun Proteomics Workflow for Protein Identification

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Shotgun quantitative label-free proteomics was performed in a nanoACQUITY UPLC system (Waters, Milford, MA, USA) coupled with a Xevo Q-TOF G2 mass spectrometer (Waters, Milford, MA, USA), as previously described [23 (link)]. Spectra were processed, and proteins were identified and quantified with Progenesis QI for Proteomics^® (Nonlinear Dynamics; Waters Corporation; version 4.0) using Apex3D (Waters) for peak detection and searching the Swiss-Prot Human proteomic database, using all the peptides for relative quantification. In order to obtain the preliminary protein dataset, the following parameters were considered: trypsin digestion with a maximum of one missed cleavage; variable modification via oxidation (M) and fixed modification via carbamidomethyl (C); false discovery rate (FDR) less than 4%; and mass error less than 20 ppm. In addition, ion-matching requirements were established to select proteins with at least one ion per peptide, three ions per protein, and one peptide per protein. Then, the final list of proteins was reduced to selected proteins identified by at least two unique peptides and proteins whose presence was detected in at least 60% of the samples.
The software CYTOSCAPE version 3.9.0 was used to build networks of molecular interactions between the identified proteins with the aid of the ClueGo and String applications.

di Flora D.C., Dionizio A., Pereira H.A., Garbieri T.F., Grizzo L.T., Dionisio T.J., Leite A.D., Silva-Costa L.C., Buzalaf N.R., Reis F.N., Pereira V.B., Rosa D.M., dos Santos C.F, & Buzalaf M.A. (2023). Analysis of Plasma Proteins Involved in Inflammation, Immune Response/Complement System, and Blood Coagulation upon Admission of COVID-19 Patients to Hospital May Help to Predict the Prognosis of the Disease. Cells, 12(12), 1601.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!