The largest database of trusted experimental protocols

Tracefinder 5

Manufactured by Thermo Fisher Scientific
Sourced in United Kingdom

TraceFinder 5.1 is a software application developed by Thermo Fisher Scientific for data processing and analysis in analytical laboratories. It provides a comprehensive suite of tools for the identification and quantification of target analytes in complex sample matrices.

Automatically generated - may contain errors

17 protocols using tracefinder 5

1

Identification of Histone Atrazine Adducts

Check if the same lab product or an alternative is used in the 5 most similar protocols
For identification of histone adducts with atrazine, the acquired UHPLC–Q-Exactive-Orbitrap-MS raw data files were converted to MGF files using Raw Converter software. LC–MS data was then preprocessed with the open-source software ProteinProspector to search for histone adducts with atrazine. Taking into consideration that under ESI ionization multicharged ions were obtained for peptides, only ions with charges of + 2 and + 3 were selected. The mass tolerance was set to 5 ppm for precursor and 10 ppm for fragment ions. Trypsin/Glu-C was specified as the cleavage enzyme and maximum missing cleavage was set at 1. Methionine oxidation was specified as variable modifications. In the “User Defined Variable Modifications” parameter, the elemental composition of C8H13N5 (179.1171 m/z) from atrazine was selected for potential adduct to amino acid residue of Cys.
For the time-dependent adduct formation study and the concentration-dependent adduct formation study, the acquired UHPLC–Q-Exactive-Orbitrap-MS raw data (in full-scan mode) was analyzed by TraceFinder 5.0 (Thermo Fisher Scientific, Mississauga, ON, Canada). The potential atrazine-modified peptide and non-modified peptide ions were selected as target ions, and the peak areas in extracted chromatograms were compared.
+ Open protocol
+ Expand
2

Targeted Metabolomics Data Processing

Check if the same lab product or an alternative is used in the 5 most similar protocols
Acquired LC-MS and GC-MS data were processed using the Thermo Scientific TraceFinder 5.0 software. Targeted metabolites were identified by matching one target and at least one confirming ion and retention time (GC-MS) or accurate mass and retention times (LC-MS) to the University of Iowa Metabolomics Core Facility’s in-house library of confirmed standards. After peak area integration by TraceFinder, NOREVA software was applied for signal drift correction on a metabolite-to-metabolite basis using the pooled QC sample that had been analyzed throughout the instrument run (16 (link)). Within individual samples, Post-NOREVA metabolite levels were divided by the total metabolite load (sum of all metabolite values) to equally weight individual metabolites for comparison across samples.
+ Open protocol
+ Expand
3

Quantification of 6-AmHap-acetamide by UPLC-PRM

Check if the same lab product or an alternative is used in the 5 most similar protocols
Quantification of 6-AmHap-acetamide
was performed in a Thermo Scientific Vanquish UPLC coupled with a
Q-Exactive Quadrupole-Orbitrap detector. The water’s HSS T3
column (2.1 mm × 100 mm, 1.8 μm particle size; Waters,
Milford, MA) and the following mobile phases were used: A (water with
10 mM NH4COOH and 0.1% HCOOH) and B (MeOH with 0.1% HCOOH).
The UPLC gradient used is described in Table S6. The column was maintained at 45 °C at a flow rate of 350 μL/min.
The injection volume was 10 μL. All data were acquired using
positive electrospray ionization (ESI) in a parallel reaction monitoring
(PRM) mode. The electrospray and source settings were as follows:
3.5 kV (capillary voltage), 320 °C (capillary temperature), 25
AU (sheath gas flow rate), 10 AU (Aux gas flow rate), and 300 °C
(Aux gas temperature). The analyte (6-AmHap-acetamide) was detected
as [M + H]+ with the PRM transition of 473.2215 > 129.0004
at 5.73 min (chromatographic retention time). Quantification was performed
using the external calibration method with a 1/X2 weighting scheme in TraceFinder 5.1 (Thermo Scientific, Waltham,
MA).
+ Open protocol
+ Expand
4

Palmitic Acid Quantification in Fungal Hyphae

Check if the same lab product or an alternative is used in the 5 most similar protocols
5 µl of each sample or palmitic acid standards in 90% acetonitrile was injected into a Vanquish LC system (Thermo Scientific, UK) using a flow rate of 0.25 mL min−1. The analytical column was Aquity UPLC CSH C18 column (1.7 um particle size, 100 mm × 2.1 mm, Waters, Manchester, UK) held at 50 °C. Starting mobile phase composition was 40% solvent B (0.1% formic acid in 95% acetonitrile/5% IPA) in A (0.1% formic acid and 80% acetonitrile in water) increasing to 54% B over 5 minutes. For column wash and equilibration the %B was increased 70–99%B over 3 minutes, held at 95% B for 2 minutes then returned to 40% B for 5 mins. Column eluant was directed in to an Orbitrap Exploris 240 mass spectrometer (ThermoFisher Scientific, UK) and ionised using electrospray ionisation in negative polarity at 2500 V. Mass measurement used full scan mode with resolution of 120,000, a m/z range of 50–500. The maximum injection time was set automatically by the software. Samples and standards were analysed in triplicate and Tracefinder 5.1 (ThermoFisher Scientific, UK) was used to construct the calibration curve and determine peak areas of palmitic acid signals in the samples. Calibration standard concentrations analysed were 1 ng, 10 ng, 100 ng, 1 µg and 10 µg. Final data were normalised per cm of hyphae per g of starting material.
+ Open protocol
+ Expand
5

Quantification of hexose sugars in fungi

Check if the same lab product or an alternative is used in the 5 most similar protocols
5 µl of each sample or the glucose and fructose standard mixture in 90% acetonitrile was injected into a Vanquish LC system (ThermoFisher Scientific, UK) using a flow rate of 0.25 mL min−1. The analytical column was Accucore-150-Amide-HILIC (2.6 µm particle size, 150 mm × 2.1 mm, ThermoFisher Scientific, UK) held at 35 °C. Starting mobile phase composition was 90% solvent B (0.1% ammonium acetate in acetonitrile) in A (0.1% ammonium acetate and water) decreasing to 60% B over 5 minutes and held constant for 4 minutes before being re-equilibrated to 90% B after 2 minutes. Column eluant was eluted into an Orbitrap Exploris 240 mass spectrometer (ThermoFisher Scientific, UK) and ionised using electrospray ionisation in negative polarity at 2500 V. Mass measurement used full scan mode resolution of 120,000, a m/z range of 5–500. The maximum injection time was set automatically by the software. Samples and standards were analysed in triplicate and Tracefinder 5.1 (ThermoFisher Scientific, UK) was used to construct the calibration curve and determine peak areas of hexose signals in the samples. Calibration standard concentrations analysed were 1 ng, 10 ng, 100 ng, 1 µg and 10 µg. Final data were normalised per cm of hyphae per g of starting material.
+ Open protocol
+ Expand
6

Quantitative Metabolomic Data Processing

Check if the same lab product or an alternative is used in the 5 most similar protocols
Areas for the 20 compounds previously selected for the model were extracted in Thermo Fisher Scientific Trace Finder 5.1 software from the.RAW files. A compound fragmentation database was obtained from Compound Discoverer from previous runs. The quan master method was employed to allow summing up of all events in cases of metabolic features eluting across multiple peaks such as cytosine and cytosine-containing compounds where source fragmentation was observed, or butrylcarnitine where multiple elution peaks were observed. The ICIS detection algorithm was employed for all metabolic features; however, other detection algorithm settings (e.g., peak detection strategy, peak threshold type) and retention times settings (i.e., detection type and RT window) were tuned individually to each metabolic feature allowing for controlled selection of the peak area in every sample. Exported areas were further processed in R for normalization and statistical analysis.
+ Open protocol
+ Expand
7

Characterization and Purification of Immunomodulatory Lipids

Check if the same lab product or an alternative is used in the 5 most similar protocols
All solvents
and reagents used in mobile phases (methanol, water, and formic acid)
were Optima LC-MS grade and were purchased from Fisher Chemicals.
DMPC, DMPG, 3D-PHAD (synthetic monophosphoryl lipid A, MPLA), and
cholesterol for liposomal preparation were purchased from Avanti Polar
Lipids Inc. and were used without further purification. Triethylamine
(Et3N) used for the hydrolysis reaction was purchased from
Sigma-Aldrich (Saint Louis, Missouri). The QS-21 working standard
was prepared from in-house HPLC-purified QS-21 purchased from the
vendor Indena (through Desert King). The QS-21 HP working standard
was generated from the purified product of the base-mediated hydrolysis
of QS-21.
ALFQ was prepared following the established procedure.31 (link) cGMP-grade ALFQ was provided by the Pilot Bioproduction
Facility (PBF) at the WRAIR.
Purification of QS-21 and QS-21
HP was done using a Shimadzu UltraFast
Liquid Chromatograph (UFLC; LC-6AD) equipped with a Shimadzu Fraction
Collector (FRC-10A). Quantitative analyses were done using a Thermo
Scientific Vanquish Flex UHPLC system coupled with a Q-Exactive Quadrupole-Orbitrap
Mass Spectrometer, controlled by Xcalibur software version 4.4. The
data were processed using Thermo Scientific TraceFinder 5.1.
+ Open protocol
+ Expand
8

UHPLC-Q-Exactive Analysis of QS-21 Saponins

Check if the same lab product or an alternative is used in the 5 most similar protocols
Quantitative analysis
of QS-21 and QS-21 HP was done using a Thermo Scientific Vanquish
UHPLC coupled with a Q-Exactive Quadrupole-Orbitrap detector. The
separation was carried out in an Agilent Zorbax Eclipse Plus C18 column
(4.6 mm ID × 50 mm, 1.8 μm particle size), using water
with 0.1% formic acid (A) and methanol with 0.1% formic acid (B) as
mobile phases with a constant flow of 0.5 mL/min at a controlled column
temperature of 35 °C. The UPLC gradient used is described in Table S5. The injection volume was set at 5 μL.
All data were acquired using negative electrospray ionization (ESI)
in parallel reaction monitoring (PRM) mode. The electrospray and source
settings were as follows: 2.5 kV (capillary voltage), 320 °C
(capillary temperature), 25 AU (sheath gas flow rate), 10 AU (Aux
gas flow rate), and 300 °C (Aux gas temperature).
Intact
QS-21 1 and QS-21 2 were detected as [M – H] with a PRM transition of m/z 1987.9169
> 485.3272 at 10.91 and 10.41 min (chromatographic RT), respectively.
QS-21 R1 and R2 derivatives eluted at 10.83 and 11.40 min, respectively,
were detected using the m/z 1855.8746
> 485.3268 PRM transition. The degradation product QS-21 HP was
detected
using a PRM transition of m/z 1511.6548
> 955.4549 at 7.19 min. Quantification was done using an external
calibration method with an equal weighting scheme in TraceFinder 5.1
(Thermo Scientific, Waltham, MA).
+ Open protocol
+ Expand
9

LC-MS Metabolomics Data Processing

Check if the same lab product or an alternative is used in the 5 most similar protocols
Acquired LC–MS data were processed using the Thermo Scientific TraceFinder 5.1 software. Targeted metabolites were identified by matching accurate mass and retention times to the University of Iowa Metabolomics Core Facility’s in-house library of confirmed standards (Table S2). During data analysis in TraceFinder, mass tolerance was set to 2 millimass units (0.002 Daltons). After peak area integration by TraceFinder, NOREVA software was applied for signal drift correction on a metabolite-to-metabolite basis using the pooled QC sample that had been analyzed throughout the instrument run as the normalizing reference [17 (link)]. We further accounted for technical variation in sample quantity and autosampler injection volume by normalizing each metabolite signal intensity to the sum of all metabolite signal intensities within a sample to generate a ratiometric metabolite fingerprint. NOREVA and ratiometrically normalized individual QC samples were superimposed on a PCA plot, providing evidence of high run quality and accurate data normalization (Figure S1). Internal standards were used to test for differences in extraction efficiency amongst samples, which were not observed. Batch effects were not present in this study since all samples were run in one batch on the LC–MS. The processing blank showed no evidence of carryover or contamination.
+ Open protocol
+ Expand
10

Quantification of Solanidine and Metabolites

Check if the same lab product or an alternative is used in the 5 most similar protocols
The serum samples had previously been analyzed on an ultra‐high performance liquid chromatography‐HRMS instrument in routine TDM of any psychiatric drugs,
14 (link) but, in this case, applied for identification and quantification of solanidine and metabolites through retrospective reprocessing of the non‐selective full scan HRMS data, as described in detail elsewhere.
8 (link),
9 (link) Solanidine and metabolites (i.e., M414, M416, and M444) were identified in the HRMS data files by accurate mass (m/z, ±5 ppm) detection at the fourth decimal and isotope ratio. The identity of solanidine was confirmed using retention time and matched tandem mass spectrometry spectrum by analyzing a reference standard purchased from Phytolab (Vestenbergsgreuth, Germany). TraceFinder 5.1 (Thermo Fisher Scientific) was used for data processing. All peaks were integrated automatically. Undetectable levels of solanidine and solanidine metabolites were truncated to the lower limit of detection (LLOD; i.e., M414, 1564; M416, 2041; M444, 1257; and solanidine, 2556) found for the corresponding analyte to enable proper calculations of metabolic ratios.
+ 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?

Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required

Sign up now

Revolutionizing how scientists
search and build protocols!