#METABOLOMICS WORKBENCH nemes_20180725_143242_mwtab DATATRACK_ID:1465 STUDY_ID:ST001028 ANALYSIS_ID:AN001686 PROJECT_ID:PR000686 VERSION 1 CREATED_ON July 30, 2018, 3:43 pm #PROJECT PR:PROJECT_TITLE Trace: Machine Learning of Signal Images for Trace-Sensitive Mass Spectrometry PR:PROJECT_TITLE – A Case Study from Single-Cell Metabolomics PR:PROJECT_TYPE Study from Single-Cell Metabolomics PR:PROJECT_SUMMARY The goal of this study was to validate the performance of a custom-written PR:PROJECT_SUMMARY software tool, called Trace, for finding molecular features from ultrasensitive PR:PROJECT_SUMMARY metabolomics experiments using high-resolution mass spectrometry. The software PR:PROJECT_SUMMARY uses a trained neural network model to extract molecular features. As model for PR:PROJECT_SUMMARY validation, we performed MS profiling of single identified cells from early PR:PROJECT_SUMMARY developing embryos of the South African clawed frog (Xenopus laevis) using a PR:PROJECT_SUMMARY custom-built capillary electrophoresis electrospray ionization platform coupled PR:PROJECT_SUMMARY to a quadrupole time-of-flight mass spectrometer. The MS dataset from these PR:PROJECT_SUMMARY measurements was manually curated for molecular features, and the resulting list PR:PROJECT_SUMMARY of molecular features were used to test the robustness and accuracy of Trace at PR:PROJECT_SUMMARY predicting molecular features that were detected from the single cells. PR:INSTITUTE University of Maryland PR:DEPARTMENT Department of Chemistry & Biochemistry PR:LABORATORY Nemes Laboratory PR:LAST_NAME Nemes PR:FIRST_NAME Peter PR:ADDRESS 0107 Chemistry Building, 8051 Regents Dr, College Park, MD 20742 PR:EMAIL nemes@umd.edu PR:PHONE 301-405-0373 PR:FUNDING_SOURCE National Cancer Institute award no. 7R03CA211635 PR:PUBLICATIONS Trace: Machine Learning of Signal Images for Trace-Sensitive Mass Spectrometry PR:PUBLICATIONS – A Case Study from Single-Cell Metabolomics #STUDY ST:STUDY_TITLE Metabolic profiling of identified single cells in Xenopus laevis embryos ST:STUDY_TYPE Metabolic profiling of single cells ST:STUDY_SUMMARY Single D11 cells were identified in 16-cell embryos of Xenopus laevis. ST:STUDY_SUMMARY Metabolites were extracted, and the extracts were analyzed using a custom-built ST:STUDY_SUMMARY capillary electrophoresis electrospray ionization platform coupled to a ST:STUDY_SUMMARY quadrupole time-of-flight mass spectrometer. The resulting metadata was analyzed ST:STUDY_SUMMARY by Trace, a custom-design software, designed to extract molecular feautres from ST:STUDY_SUMMARY trace-sensitive metabolomics experiments. The results were validated against ST:STUDY_SUMMARY molecular features that were extracted by manual curation of the same raw mass ST:STUDY_SUMMARY spectrometer files. ST:INSTITUTE University of Maryland ST:DEPARTMENT Department of Chemistry & Biochemistry ST:LABORATORY Nemes Laboratory ST:LAST_NAME Nemes ST:FIRST_NAME Peter ST:ADDRESS 0107 Chemistry Building, 8051 Regents Dr, College Park, MD 20742 ST:EMAIL nemes@umd.edu ST:PHONE 3014050373 ST:NUM_GROUPS 5 biological replicates (different cells from different embryos) + 1-to-3 ST:NUM_GROUPS technical replicates (same extract analyzed multiple times) ST:TOTAL_SUBJECTS 5 different D11 cells were analyzed, each from a different embryo ST:PUBLICATIONS Trace: Machine Learning of Signal Images for Trace-Sensitive Mass Spectrometry ST:PUBLICATIONS – A Case Study from Single-Cell Metabolomics #SUBJECT SU:SUBJECT_TYPE Other SU:SUBJECT_SPECIES Xenopus laevis SU:TAXONOMY_ID 8355 SU:AGE_OR_AGE_RANGE Embryos were obtained from natural mating of frogs (Nasco) SU:WEIGHT_OR_WEIGHT_RANGE Sexually mature male and female frogs SU:GENDER Not applicable #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS 2016-03-15_EP03 D11 cell-E2-2 D11cellE2T2 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-03-15_EP03 D11 cell-E2-1 D11cellE2T1 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-03-14_EP04 D11 cell-E1-1 D11cellE4T1 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-02-17_RM02 v23 D11 D11cellE1T1 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-06-01_EP05 u-cap E3-D11-1 D11cellE3T1 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-06-01_EP09 u-cap E3-D11-2 D11cellE3T2 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-06-01_EP11 u-cap E3-D11-3 D11cellE3T3 Embryo Type:WT SUBJECT_SAMPLE_FACTORS 2016-06-02_EP04 u-cap E1-D11-1 D11cellE5T1 Embryo Type:WT #COLLECTION CO:COLLECTION_SUMMARY Cells were identified based on morphology, pigmentation, and location in the CO:COLLECTION_SUMMARY embryo in comparision to established cell-fate maps for 16-cell Xenopus laevis CO:COLLECTION_SUMMARY embryos. A portion of the identified D11 cell was microaspirated using a CO:COLLECTION_SUMMARY fabricated microcapillary. CO:COLLECTION_PROTOCOL_ID Liu 2018 Metabolomics Workbench Protocol.pdf CO:SAMPLE_TYPE Embryonic cells CO:COLLECTION_METHOD Microaspiration of cell content CO:COLLECTION_FREQUENCY 1 collection per cell CO:COLLECTION_DURATION 5 s for aspiration CO:VOLUMEORAMOUNT_COLLECTED Ca. 10 nL per aspiration CO:STORAGE_CONDITIONS -80℃ CO:COLLECTION_TUBE_TEMP chilled on ice #TREATMENT TR:TREATMENT_SUMMARY All protocols related to the handling and manipulation of animals were approved TR:TREATMENT_SUMMARY by the Institutional Animal Care and Use Committee (IACUC) of the George TR:TREATMENT_SUMMARY Washington University (Washington, DC) and the University of Maryland, College TR:TREATMENT_SUMMARY Park (College Park, MD). TR:TREATMENT_PROTOCOL_ID IACUC #A311 (George Washington University) and IACUC #R-DEC-17-57 (University of TR:TREATMENT_PROTOCOL_ID Maryland, College Park) TR:TREATMENT_PROTOCOL_FILENAME Liu 2018 Metabolomics Workbench Protocol.pdf TR:TREATMENT_PROTOCOL_COMMENTS Embryos were dejellied using 2% cystine solution, cultured in 100% Steinberg's TR:TREATMENT_PROTOCOL_COMMENTS solution (media), and used without further treatment. #SAMPLEPREP SP:SAMPLEPREP_SUMMARY Ca. 10 nL of cell content were aspirated from identified cells. The aspirated SP:SAMPLEPREP_SUMMARY material was ejected into ~4 uL of aqueous mixture of 40% acetonitrile and 40% SP:SAMPLEPREP_SUMMARY methanol to extract metabolites. SP:SAMPLEPREP_PROTOCOL_ID Cold aqueous acetonitrile-methanold extraction SP:SAMPLEPREP_PROTOCOL_FILENAME Liu 2018 Metabolomics Workbench Protocol.pdf SP:PROCESSING_STORAGE_CONDITIONS On ice SP:EXTRACTION_METHOD In cold aqueous mixture of 40% acetonitrile and 40% methanol. SP:EXTRACT_STORAGE -80℃ SP:SAMPLE_RESUSPENSION None. The cells were stored in the extraction solution. The extract was not SP:SAMPLE_RESUSPENSION dried or processed further. SP:SUBCELLULAR_LOCATION Unknown #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE CE CH:INSTRUMENT_NAME Custom built CE system CH:COLUMN_NAME Bare fused silica capillary #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME Nemes Laboratory AN:OPERATOR_NAME Erika Portero AN:SOFTWARE_VERSION Compass ver. 4.3 AN:ACQUISITION_DATE 2016 Feb-to-June AN:DATA_FORMAT .d (Bruker) #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Bruker Impact HD MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE POSITIVE MS:ACQUISITION_DATE 2016 Feb-to-June MS:CAPILLARY_TEMPERATURE Ca. 100 degC MS:CAPILLARY_VOLTAGE -1700 V MS:DRY_GAS_FLOW 2 L/min MS:DRY_GAS_TEMP 100 degC MS:ION_SOURCE_TEMPERATURE 100 degC MS:MASS_ACCURACY <10 ppm MS:RESOLUTION_SETTING ~45000 FWHM MS:SCANNING_CYCLE 2 Hz spectral MS:SCANNING_RANGE m/z 50-550 MS:MS_RESULTS_FILE ST001028_AN001686_Results.txt UNITS:PeakAreaInCounts #END