#METABOLOMICS WORKBENCH araskind_20150605_9931401_mwtab.txt DATATRACK_ID:271 STUDY_ID:ST000174 ANALYSIS_ID:AN000268 PROJECT_ID:PR000151 VERSION 1 CREATED_ON 2016-09-17 #PROJECT PR:PROJECT_TITLE Nlrp3 PR:PROJECT_SUMMARY Unstimulated vs Stimulated n=4 PR:INSTITUTE University of Michigan PR:DEPARTMENT Pathology PR:LABORATORY Nunez PR:LAST_NAME Nunez PR:FIRST_NAME Gabriel PR:ADDRESS Ann Arbor, MI PR:EMAIL bclx@med.umich.edu PR:PHONE 734-764-8514 #STUDY ST:STUDY_TITLE Mouse macrophages from bone marrow treatment ST:STUDY_TYPE See attached document ST:STUDY_SUMMARY The experiment was done on 4/2/2014 and consists of 7 experimental conditions x ST:STUDY_SUMMARY replicates per condition = 14 samples. The experimental duplicates are numbered ST:STUDY_SUMMARY (i.e. 1 and 2, 3 and 4, etc.). All samples are primary mouse macrophages ST:STUDY_SUMMARY from bone marrow. 12 million cells were plated in each 10 cm tissue culture ST:STUDY_SUMMARY plate (Corning) and the following day the cells were stimulated with LPS, ST:STUDY_SUMMARY CL097 and/or Gardiquimod in 10 mls of 2% FBS 25 mM HEPES IMDM. After 60 minutes ST:STUDY_SUMMARY medium was aspirated and the cells were rapidly wahsed with 15 mls of 150 mM ST:STUDY_SUMMARY acetate and after aspiration liquid nitrogen was poured on top of the cells and ST:STUDY_SUMMARY to -80 C freezer. Sample pairs 7 - 8 and 9-10 were treated identically except ST:STUDY_SUMMARY the rinse was done with ammonium acetate (7 and 8) or distilled water (9 and ST:STUDY_SUMMARY to evaluate the effect of the rinse solution in the quality of the obtained ST:INSTITUTE University of Michigan ST:DEPARTMENT Biomedical Research Core Facilities ST:LABORATORY Metabolomics core ST:LAST_NAME Kachman ST:FIRST_NAME Maureen ST:ADDRESS 6300 Brehm Tower, 1000 Wall Street, Ann Arbor, MI 48105-5714 ST:EMAIL mkachman@umich.edu ST:PHONE - ST:NUM_GROUPS 10 ST:TOTAL_SUBJECTS 14 #SUBJECT SU:SUBJECT_TYPE - SU:SUBJECT_SPECIES Mus musculus SU:TAXONOMY_ID 10090 #SUBJECT_SAMPLE_FACTORS: SUBJECT(optional)[tab]SAMPLE[tab]FACTORS(NAME:VALUE pairs separated by |)[tab]Additional sample data SUBJECT_SAMPLE_FACTORS SU0003757 S00014354 Agonist:CL097 | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003757 S00014355 Agonist:CL097 | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003758 S00014356 Agonist:Gardiquimod | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003758 S00014357 Agonist:Gardiquimod | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003756 S00014352 Agonist:Gramicidin | Priming:LPS | Rinse:H2O rinse SUBJECT_SAMPLE_FACTORS SU0003755 S00014350 Agonist:Gramicidin | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003754 S00014348 Agonist:Gramicidin | Priming:None | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003756 S00014353 Agonist:Gramidicin | Priming:LPS | Rinse:H2O rinse SUBJECT_SAMPLE_FACTORS SU0003755 S00014351 Agonist:Gramidicin | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003754 S00014349 Agonist:Gramidicin | Priming:None | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003753 S00014346 Agonist:None | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003753 S00014347 Agonist:None | Priming:LPS | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003752 S00014344 Agonist:None | Priming:None | Rinse:None SUBJECT_SAMPLE_FACTORS SU0003752 S00014345 Agonist:None | Priming:None | Rinse:None #COLLECTION CO:COLLECTION_SUMMARY - CO:SAMPLE_TYPE Cells #TREATMENT TR:TREATMENT_SUMMARY - #SAMPLEPREP SP:SAMPLEPREP_SUMMARY - SP:SAMPLEPREP_PROTOCOL_FILENAME Gly-TCA-nucleotides_analysis_protocol-2015-03-09.docx #CHROMATOGRAPHY CH:CHROMATOGRAPHY_TYPE HILIC CH:INSTRUMENT_NAME Agilent 1260 CH:COLUMN_NAME Phenomenex Luna NH2 (150 x 1mm, 3um) CH:METHODS_ID AQM020 CH:METHODS_FILENAME QTOF-002-HILIC-35min-1mm-No_Insert.m.zip #ANALYSIS AN:ANALYSIS_TYPE MS AN:LABORATORY_NAME MRC2 (University of Michigan) AN:ACQUISITION_PARAMETERS_FILE QTOF-002-HILIC-35min-1mm-No_Insert.m.zip AN:PROCESSING_PARAMETERS_FILE EX00289-MassHunterQuant-GlyTCA-LCMS-DataAnalysis-Method.m.zip #MS MS:MS_COMMENTS - MS:INSTRUMENT_NAME Agilent 6530 QTOF MS:INSTRUMENT_TYPE QTOF MS:MS_TYPE ESI MS:ION_MODE NEGATIVE #MS_METABOLITE_DATA MS_METABOLITE_DATA:UNITS pmol/mg protein MS_METABOLITE_DATA_START Samples S00014354 S00014355 S00014356 S00014357 S00014352 S00014350 S00014348 S00014353 S00014351 S00014349 S00014346 S00014347 S00014344 S00014345 Factors Agonist:CL097 | Priming:LPS | Rinse:None Agonist:CL097 | Priming:LPS | Rinse:None Agonist:Gardiquimod | Priming:LPS | Rinse:None Agonist:Gardiquimod | Priming:LPS | Rinse:None Agonist:Gramicidin | Priming:LPS | Rinse:H2O rinse Agonist:Gramicidin | Priming:LPS | Rinse:None Agonist:Gramicidin | Priming:None | Rinse:None Agonist:Gramidicin | Priming:LPS | Rinse:H2O rinse Agonist:Gramidicin | Priming:LPS | Rinse:None Agonist:Gramidicin | Priming:None | Rinse:None Agonist:None | Priming:LPS | Rinse:None Agonist:None | Priming:LPS | Rinse:None Agonist:None | Priming:None | Rinse:None Agonist:None | Priming:None | Rinse:None 2 or 3-phosphoglycerate_2PG/3PG 1.0494 1.3942 2.8205 2.5683 4.1072 10.2820 3.7042 3.2632 3.9296 3.7430 3.8949 8.1576 7.9188 3.8938 6-Phosphogluconate 0.5471 0.8237 1.4006 0.9114 1.4036 6.0292 1.3281 2.8558 7.5143 1.0025 1.5099 8.3502 3.5302 1.7132 Acetyl-CoA 0.5690 0.2568 0.6756 0.6233 1.3706 0.2415 0.5077 0.1319 0.4947 0.6204 2.1008 0.6814 3.0192 1.9674 ADP 52.6842 57.5349 44.8088 26.1730 21.5733 13.2795 74.4780 4.2287 65.5902 54.8321 55.7849 161.8897 205.1254 82.6191 alpha-Ketoglutarate 47.0842 128.0396 28.0642 7.6897 9.7954 168.6839 42.3369 74.9946 105.6329 19.5165 18.9061 329.3616 83.1426 35.2645 AMP 76.5378 27.1297 159.0582 194.3999 241.4281 16.8205 230.5189 5.8977 77.3951 269.3636 397.6437 171.0569 906.6525 506.6701 ATP 32.8171 41.8404 117.6068 120.9454 71.8726 120.2651 87.3411 38.0911 75.9822 80.3558 260.3726 440.6007 532.2357 277.6291 Citrate or Isocitrate 2.6971 3.7767 6.4447 6.1458 12.9459 22.5954 11.6099 5.4284 11.9180 10.6722 11.8280 26.1497 38.1070 16.1228 flavin adenine dinucleotide 3.5169 6.6631 18.6692 19.2117 28.5602 35.7964 36.9917 13.2146 27.1073 32.4625 34.3196 67.6484 83.7510 47.9034 Fructose 1_6-bisphosphate 652.7995 723.0678 518.3133 486.8363 985.7306 934.8419 956.4265 565.3154 627.4335 848.1710 1116.8519 1949.5882 3464.2210 2146.2010 Glucose 6-phosphate or fructose-6-phosphate 1.9257 2.2267 2.0749 2.0944 1.1311 3.1031 2.1799 0.5065 2.0742 2.2151 3.6277 7.4183 9.3177 4.0424 Malate 55.1973 77.9213 32.9770 34.6275 23.6834 85.6886 42.0895 18.8198 37.5535 41.1506 149.2816 209.2503 304.9810 167.0775 Nicotinamide Adenine Dinucleotide oxidized 23.1419 29.5281 34.8950 35.0931 25.6436 47.9136 37.1797 13.7641 31.1558 35.7789 70.2599 110.2385 176.0899 80.1814 nicotinamide adenine dinucleotide phosphate oxidized 3.3111 5.9421 3.7421 2.9606 2.0484 9.8581 5.3459 3.5196 5.0331 4.2106 3.1993 20.9558 10.9782 3.8073 nicotinamide adenine dinucleotide phosphate reduced 18.1414 1.8881 34.6331 39.1564 83.2599 4.1868 15.7800 3.6109 19.9811 29.4112 170.7335 16.0425 242.3769 144.0608 Nicotinamide Adenine Dinucleotide reduced 35.8611 23.7141 27.2596 29.1568 14.6299 31.9921 20.5127 4.8158 28.4318 25.4223 59.2030 82.3655 121.8218 69.1843 Phosphoenolpyruvate 0.3489 0.3971 1.2025 1.1612 0.3948 2.4869 1.6574 0.2631 1.8369 1.5782 2.4143 3.2765 4.4455 2.6023 Sucrose 4.2925 5.6026 10.9184 10.4761 30.0090 34.3012 22.8583 18.5218 19.1234 20.8855 32.8368 48.1953 62.7565 35.6370 MS_METABOLITE_DATA_END #METABOLITES METABOLITES_START metabolite_name moverz_quant ri ri_type pubchem_id inchi_key kegg_id other_id other_id_type 2 or 3-phosphoglycerate_2PG/3PG 724 2PG/3PG UM_Target_ID 6-Phosphogluconate 91493 6PG UM_Target_ID Acetyl-CoA 444493 aCoA UM_Target_ID ADP 6022 ADP UM_Target_ID alpha-Ketoglutarate 51 AKG UM_Target_ID AMP 6083 AMP UM_Target_ID ATP 5957 ATP UM_Target_ID Citrate or Isocitrate 311 CIT/ICIT UM_Target_ID flavin adenine dinucleotide 46906035 FAD UM_Target_ID Fructose 1,6-bisphosphate 21125049 FBP UM_Target_ID Glucose 6-phosphate or fructose-6-phosphate 5958 G6P/F6P UM_Target_ID Malate 525 MAL UM_Target_ID Nicotinamide Adenine Dinucleotide oxidized 15938971 NAD UM_Target_ID nicotinamide adenine dinucleotide phosphate oxidized 15938972 NADP UM_Target_ID nicotinamide adenine dinucleotide phosphate reduced 22833512 NADPH UM_Target_ID Nicotinamide Adenine Dinucleotide reduced 439153 NADH UM_Target_ID Phosphoenolpyruvate 1005 PEP UM_Target_ID Sucrose 5988 SUC UM_Target_ID METABOLITES_END #END