Proteins were tryptic digested before passing through the C18 column packed with 1.9umAqua C18 material (Phenomenex). Peptides eluted from the LC column were directly electrosprayed into a Q-Exactive plus mass spectrometer (ThermoFisher, San Jose, CA). MS scan functions and HPLC solvent gradients were controlled by the Xcalibur data system (ThermoFisher). All raw files were processed using Maxquant software (version 1.5.3.30) (Beer et al. 2017 (link)) for feature detection, database searching, and protein/peptide quantification. MS/MS spectra were searched against the UniProtKB/Swiss-Prot human database (containing 20,386 reviewed sequences). The minimum peptide length was seven amino acids, and maximum peptide mass was 4600 Da. The allowed missed cleavages for each peptide was two. The second peptide search was activated to identify coeluting and cofragmented peptides from one MS/MS spectrum. Both peptides and proteins were filtered with a maximum FDR of 0.01. LFQ calculations were performed separately in each parameter group. Both unique and razor peptides were selected for protein quantification. Other unmentioned parameters were the default settings of the Maxquant software (Beer et al. 2017 (link)).
Xcalibur data system
Manufactured by Thermo Fisher Scientific
Sourced in United States, Italy
The Xcalibur data system is a comprehensive software solution designed for data acquisition, processing, and management in analytical laboratories. It provides a streamlined interface for controlling and monitoring various Thermo Fisher Scientific mass spectrometry instruments.
Automatically generated - may contain errors
Lab products found in correlation
Dionex ultimate 3000 uplc, by Thermo Fisher Scientific (6 mentions)
Easy spray source, by Thermo Fisher Scientific (4 mentions)
Linear ion trap ltq xl, by Thermo Fisher Scientific (4 mentions)
Ltq xl linear ion trap mass spectrometer, by Thermo Fisher Scientific (3 mentions)
Dionex ultimate 3000, by Thermo Fisher Scientific (2 mentions)
Accela pda detector, by Thermo Fisher Scientific (2 mentions)
Ltq xl mass spectrometer, by Thermo Fisher Scientific (2 mentions)
Lcq fleet ion trap mass spectrometer, by Thermo Fisher Scientific (2 mentions)
Q exactive plus mass spectrometer, by Thermo Fisher Scientific (1 mentions)
Pierce ms compatible magnetic ip kit, by Thermo Fisher Scientific (1 mentions)
Thermo fusion tribrid mass spectrometer, by Thermo Fisher Scientific (1 mentions)
Trypsin gold, by Promega (1 mentions)
Spherisorb s3 ods 2 c18, by Waters Corporation (1 mentions)
Accela 1250 pump, by Thermo Fisher Scientific (1 mentions)
Ultra clear system, by Siemens (1 mentions)
Hypersil gold aq, by Thermo Fisher Scientific (1 mentions)
Q exactive hybrid quadrupole orbitrap mass spectrometer, by Thermo Fisher Scientific (1 mentions)
Esi source, by Thermo Fisher Scientific (1 mentions)
Tmt 6 plex reagent, by Thermo Fisher Scientific (1 mentions)
Ltq orbitrap velos, by Thermo Fisher Scientific (1 mentions)
38 protocols using xcalibur data system
1
Quantitative Proteomic Analysis Pipeline
2
Immunoprecipitation and Mass Spectrometry Analysis
3
HPLC-DAD-ESI/MSn Profiling of Phenolic Compounds
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The obtained extracts were re-dissolved in 20% aqueous ethanol at 10 mg/mL, filtered through a 0.22 µm nylon syringe filter, and analyzed by HPLC-DAD-ESI/MSn in a Dionex Ultimate 3000 UPLC system (Thermo Scientific, San Jose, CA, USA). The equipment consisted of a diode array detector coupled to an electrospray ionization mass detector, a quaternary pump, an auto-sampler (kept at 5 °C), a degasser, and an automated thermostated column section (kept at 35 °C). A Waters Spherisorb S3 ODS-2 C18 (3 µm, 4.6 mm × 150 mm, Waters, Milford, MA, USA) column was used. Data were collected simultaneously with DAD and in negative mode detection on a Linear Ion Trap LTQXL mass spectrometer (Thermo Scientific, San Jose, CA, USA), following a previously described procedure [22 (link)]. An Xcalibur® data system (Thermo Scientific, San Jose, CA, USA) was used in data acquisition. Phenolic compounds were identified by comparing their retention times and UV–Vis and mass spectra with standards, when available. Otherwise, compounds were identified tentatively by comparing data obtained from available information published in the literature. For quantitative analysis, calibration curves obtained from available standards or closely related standards were used. The results were expressed as mg/g of extract.
4
Peptide Identification via Tandem MS
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Peptides eluted from the microcapillary
column were electrosprayed directly into an LTQ-XL mass spectrometer
(Thermo Fisher, USA) with the application of a distal 2.4 kV spray
voltage. A cycle of one full-scan mass spectrum (300–2000 m/z) followed by five data-dependent MS/MS
spectra at a 35% normalized collision energy was repeated continuously
throughout each step of the multidimensional separation. To prevent
repetitive analysis, dynamic exclusion was enabled with a repeat count
of 1, a repeat duration of 30 s, and an exclusion list size of 200.
Application of mass spectrometer scan functions and HPLC solvent gradients
were controlled by the Xcalibur data system (Thermo, USA).
column were electrosprayed directly into an LTQ-XL mass spectrometer
(Thermo Fisher, USA) with the application of a distal 2.4 kV spray
voltage. A cycle of one full-scan mass spectrum (300–2000 m/z) followed by five data-dependent MS/MS
spectra at a 35% normalized collision energy was repeated continuously
throughout each step of the multidimensional separation. To prevent
repetitive analysis, dynamic exclusion was enabled with a repeat count
of 1, a repeat duration of 30 s, and an exclusion list size of 200.
Application of mass spectrometer scan functions and HPLC solvent gradients
were controlled by the Xcalibur data system (Thermo, USA).
5
HPLC-HRMS Enantioselective Analysis of Praziquantel
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The HPLC–HRMS chromatographic system included an Accela 1250 pump, a PAL HTC autosampler, an Accela PDA detector and a Xcalibur data system (ThermoFisher Scientific, Waltham, MA, USA). Water was purified by an Ultra Clear system (Siemens, Berlin, Germany). Chromatographic separation of R-PZQ, S-PZQ and IS mebendazole were obtained on a column of Daicel CHIRALPAK® AS-RH (4.6 mm × 150 mm, 5 μm) fitted with a guard column (10 × 4.0 mm, 3 μm) at room temperature (about 25 °C). The autosampler was set at 4 °C during analysis. The mobile phase consisted of methanol and formic acid (90:10, v/v) at the flow rate of 0.55 ml/min. The relative parameters were as follows: ESI ion source, 3.00 kV spray voltage, sheath gas (N2) and auxiliary gas (N2) 35 Arb and 10 Arb, respectively, capillary heater temperature 320 °C. The [M -H]+ quasi-molecular ion peaks of R-PZQ and S-PZQ were detected at 313.1911 mass-to-charge (m/z) ratio (the two enantiomers have the same molecular weight and molecular structure and differ only with regard to symmetry), while the IS (mebendazole) was detected at 296.1036 m/z. The total run time was 25.0 min.
6
Quantifying Brassinosteroids in Rice Panicles
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
At the SPD and PMC stages, 150 young panicles were collected from each plot to measure the concentration of BRs. The extraction and purification of BRs from the panicles were based on the method with modification (Zhang et al., 2020 (link)). Fresh panicles were frozen in liquid N, ground into a fine powder, transferred (0.5–0.8 g) to a 10-mL centrifuge tube, added 4 mL of acetonitrile for extraction, and refrigerated overnight at −20°C. Extraction, dehydration, and double layer-solid phase extraction (DL/SPE) were performed following the described method (Chen et al., 2009 (link)). The BRs were quantified using high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS). The levels of 24-epicastasterone (24-epiCS) and 28-homobrassinolide (28-homoBL) were measured, as they are the most important BRs in rice plants and exhibit high biological activity. Quantification (pmol g−1 fresh weight) was by the ratio of the total area converted based on the multiple reaction monitoring (MRM) of BRs, with a calibration by corresponding standard BRs. The Xcalibur data system (Thermo Fisher Scientific, Waltham, MA, USA) was used for data collection and analysis.
7
UHPLC-MS Compound Identification Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The UHPLC was coupled to an LTQ XL Linear Ion Trap 2D mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA), equipped with an orthogonal electrospray ionization source operating in negative mode. The nitrogen sheath and auxiliary gas were 50 and 10 (arbitrary units), respectively. The spray voltage was 5 kV and the capillary temperature was 275 °C. The capillary and tune lens voltages were set at −28 V and −115 V, respectively. The data acquisition was carried out by using Xcalibur® data system (Thermo Scientific, San Jose, CA, USA). The identification of compounds obtained by chemical analysis of the crude extract was carried out by comparing their chromatographic characteristics (e.g., retention time, UV-Vis maximum absorption (λmax, 190–700 nm), and mass spectrum (m/z)) with data available in the literature.
8
Isocratic LC-MS Analysis of Small Molecules
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The analysis was performed with a Wadose LC isocratic pump, interfaced with a Q-Exactive Hybrid Quadrupole-Orbitrap mass spectrometer equipped with an ESI source (Thermo Fisher Scientific, Waltham, MA, USA). The MS functions were controlled by the Xcalibur data system (Thermo Fisher Scientific), whereas injection and HPLC solvent elution were monitored and controlled by our software developed on LabVIEW. The analytical column was a semi-polar Hypersil Gold aQ (50 × 1 mm, 1.9 µm, 175 Å, Thermo Fisher Scientific). The mobile phase consisted of acetonitrile (ACN)–0.1% formic acid and water–0.1% formic acid. Elution was performed with 10% and 20% ACN at a constant flow rate of 110 µL·min−1. Experiments were conducted at 40 °C.
9
Quantitative Proteomic Analysis of ES2 Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Three biological replicates of lysates of ES2 cells treated with C43 and AC220 for 16 h (MG132 for the last 4 h) were precipitated by methanol/chloroform, and protein samples were resuspended in 8 M urea/100 mM TEAB, pH 8.0, and reduced, then alkylated with 10 mM Tris (2-carboxyethyl) phosphine hydrochloride and 55 mM iodoacetamide, followed by tryosin digestion. The resultant peptides were labeled with 6-plex TMT reagents as the user manual describes (Thermo Fisher Scientific). The resulted 6-plex mixture of peptides was analyzed on a hybrid LTQ-Orbitrap Velos mass spectrometer (Thermo Fisher Scientific) using a modified 12-step MudPIT separation that has been described previously (Washburn et al., 2001 (link); Rauniyar et al., 2015 (link)). Application of ms scan functions and HPLC solvent gradients were controlled with the Xcalibur data system (Thermo Fisher Scientific).
10
Multidimensional Peptide Separation and Mass Spectrometry
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Peptides
eluted from
the microcapillary column were electrosprayed directly into an LTQ-XL
mass spectrometer (Thermo Finnigan, Palo Alto, CA) with the application
of a distal 2.4 kV spray voltage. A cycle of one full-scan mass spectrum
(300–2000 m/z) followed by
five data-dependent MS/MS spectra at a 35% normalized collision energy
was repeated continuously throughout each step of the multidimensional
separation. To prevent repetitive analysis, dynamic exclusion was
enabled with a repeat count of 1, a repeat duration of 30 s, and an
exclusion list size of 200. Application of mass spectrometer scan
functions and HPLC solvent gradients was controlled by the Xcalibur
data system (Thermo, San Jose, CA).
eluted from
the microcapillary column were electrosprayed directly into an LTQ-XL
mass spectrometer (Thermo Finnigan, Palo Alto, CA) with the application
of a distal 2.4 kV spray voltage. A cycle of one full-scan mass spectrum
(300–2000 m/z) followed by
five data-dependent MS/MS spectra at a 35% normalized collision energy
was repeated continuously throughout each step of the multidimensional
separation. To prevent repetitive analysis, dynamic exclusion was
enabled with a repeat count of 1, a repeat duration of 30 s, and an
exclusion list size of 200. Application of mass spectrometer scan
functions and HPLC solvent gradients was controlled by the Xcalibur
data system (Thermo, San Jose, CA).
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?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!