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| Status |
Public on Aug 23, 2011 |
| Title |
Uninfected 2 |
| Sample type |
other |
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| Source name |
Gallbladder
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
gender: Female
infection status: uninfected
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| Treatment protocol |
Oral infection with 3-8x10^7 Salmonella enterica serovar Typhimurium
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| Extracted molecule |
other |
| Extraction protocol |
To extract metabolites from gallbladders, 2 ml of chloroform:methanol (2:1) was added to each sample. Tissues were then homogenized using a tungsten bead and a Mixer Mill MM 301 (Retsch, Haan, Germany). The samples were then transferred to ice, and 1 ml of cold H2O was added. The samples were centrifuged at 3250 x g for 10 minutes at 4°C. The upper phase was discarded, and another 500 µl of cold H2O was added. The samples were centrifuged again at 3250 x g for 10 minutes at 4°C. The upper phase was discarded, and the lower phase was transferred to a new glass tube and dried under N2 gas. All extracts were kept at -80˚C until further use.
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| Label |
Not applicable
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| Label protocol |
Not applicable
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| Hybridization protocol |
Not applicable
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| Scan protocol |
Not applicable
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| Description |
Metabolites
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| Data processing |
Raw mass spectrometry data were processed using a custom-developed software package, as described elsewhere (Han et al. 2008. Metabolomics. 4:128-140). Mass spectra were batch-processed using a custom VBA script, within the instrument vendor’s data processing software package, DataAnalysis®. First, all the raw mass spectra acquired in positive and negative ion modes were internally calibrated with the reference masses of the spiked standard compounds and a ubiquitous known contaminant, N-butylbenzensulfonamide. Monoisotopic peaks corresponding to the isotopic distribution patterns were then automatically determined and those with signal-to-noise ratios above 3 were picked. The m/z values were subsequently converted to neutral masses by subtracting 1.007276 for positive ion mode or adding 1.007276 for negative ion mode. Next, the resulting mass lists from the individual mass spectra within each paired sample group were further processed with another custom software program developed with LabVIEW® (National Instruments, Austin, USA). The first step of this program is to remove the adduct ions, e.g., (M+Na)+ and/or (M+K)+ in positive ion mode and (M+Cl)- in negative ion mode, from the mass lists based on the expected mass differences for these ions within 2 ppm. Next, the peak intensity of each unique monoisotopic neutral mass was normalized to the intra-spectrum total ion intensity. Masses observed in at least 50% of the spectra from each paired sample group were aligned into unique masses (metabolite features) from the masses that matched within 2 ppm. Finally, a two-dimensional data matrix (neutral mass vs. peak intensity) was generated for each group and saved in a tab-delimited text format. To detect metabolite differences between the different sample groups, the masses were filtered in each list for metabolites that were present on one set of samples, but not the other. Additionally, the intensities were averaged of each unique mass in each set of biological replicates and calculated the ratio between the sample groups. To assign possible metabolite identities to the masses present only in one set or showing at least 2-fold changes, the monoisotopic neutral masses of interest were queried against MassTrix (http:// masstrix.org), software designed to incorporate mass queries into metabolic pathways [21]. Masses were searched against the Mus musculus database within an error of 3 ppm.
Masses (daltons) with relative intensity are provided in the "masses_and_intensities.txt" file on the Series record. TAR archive of raw spectral files and a readme with Sample associations are also provided there.
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| Submission date |
Mar 01, 2011 |
| Last update date |
Aug 23, 2011 |
| Contact name |
L. Caetano M. Antunes |
| E-mail(s) |
antunes@mail.ubc.ca
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| Phone |
604-827-3921
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| Organization name |
The University of British Columbia
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| Department |
Michael Smith Laboratories
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| Lab |
Finlay Lab
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| Street address |
#367 - 2185 East Mall
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| City |
Vancouver |
| State/province |
BC |
| ZIP/Postal code |
V6T 1Z4 |
| Country |
Canada |
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| Platform ID |
GPL10454 |
| Series (1) |
| GSE27604 |
A metabolomics analysis of gallbladder lipid droplet formation due to Salmonella infection |
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| Supplementary data files not provided |
| Processed data are available on Series record |
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