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MB Sample ID: SA300467
| Local Sample ID: | T14_253R |
| Subject ID: | SU002904 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002904 |
| Subject Type: | Human |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| T14_253R | SA300467 | FL036474 | TTTS | Group |
Collection:
| Collection ID: | CO002897 |
| Collection Summary: | Amniotic fluid was previously collected and banked from pregnant individuals with monochorionic-diamniotic twins complicated by TTTS undergoing fetoscopic laser ablation of placental anastomoses at the UTHealth Houston Fetal Center at Children’s Memorial Hermann Hospital. Samples were collected from the recipient twin sac immediately upon entry with the operative cannula and prior to placental laser ablation or amnioinfusion for improving the visualization. The amniotic fluid was centrifuged, and the supernatant was stored at -80 C for future use. Frozen genetic amniocentesis samples, discarded from further analyses, served as controls. For TTTS cases, demographics, clinical characteristics, and outcomes were abstracted from the Fetal Center research database which is maintained by trained research staff. For controls, available clinical variables were limited to maternal age, gestational age, indication for amniocentesis, and genetic testing results. |
| Sample Type: | Amniotic fluid |
Treatment:
| Treatment ID: | TR002913 |
| Treatment Summary: | not applicable |
Sample Preparation:
| Sampleprep ID: | SP002910 |
| Sampleprep Summary: | Amniotic fluid metabolites were extracted by addition of 1 part amniotic fluid to 15 parts 70% acetonitrile in ddH2O (vol:vol). The mixture was briefly vortexed and then centrifuged for 5 min at 16,000 × g to pellet precipitated proteins. The protein pellet was solubilized in 0.2M NaOH and quantified by DC Protein Assay (Bio-Rad). The volume of metabolite extract was normalized by amniotic protein content. |
Combined analysis:
| Analysis ID | AN004550 | AN004551 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Normal phase | Normal phase |
| Chromatography system | Agilent 6550 QTOF | Agilent 6550 QTOF |
| Column | MicroSolv Diamond Hydride (150 x 2.1mm, 4um) | MicroSolv Diamond Hydride (150 x 2.1mm, 4um) |
| MS Type | ESI | ESI |
| MS instrument type | QTOF | QTOF |
| MS instrument name | Agilent 6550 QTOF | Agilent 6550 QTOF |
| Ion Mode | POSITIVE | NEGATIVE |
| Units | intensity | intensity |
Chromatography:
| Chromatography ID: | CH003419 |
| Instrument Name: | Agilent 6550 QTOF |
| Column Name: | MicroSolv Diamond Hydride (150 x 2.1mm, 4um) |
| Column Temperature: | 25 |
| Flow Gradient: | 0-1.0 min, 99% B; 1.0-15.0 min, to 20% B; 15.0 to 29.0, 0% B; 29.1 to 37min, 99% B |
| Flow Rate: | 0.4 mL/min |
| Solvent A: | 50% isopropanol/50% water; 0.025% acetic acid |
| Solvent B: | 90% acetonitrile/10% water; 5 mM ammonium acetate |
| Chromatography Type: | Normal phase |
MS:
| MS ID: | MS004297 |
| Analysis ID: | AN004550 |
| Instrument Name: | Agilent 6550 QTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | MS acquisition comments: For aqueous normal phase LC/MS-based metabolomics, amniotic fluid metabolites were extracted by addition of 1 part amniotic fluid to 15 parts 70% acetonitrile in ddH2O (vol:vol). The mixture was briefly vortexed and then centrifuged for 5 min at 16,000 × g to pellet precipitated proteins. The protein pellet was solubilized in 0.2M NaOH and quantified by DC Protein Assay (Bio-Rad). The volume of metabolite extract was normalized by amniotic protein content. An aliquot of the resulting extract (3 mL) was subjected to LC/MS untargeted metabolite profiling in positive and negative ion modes as described previously described,64 using a platform comprised of an Agilent Model 1290 Infinity II liquid chromatography system coupled to an Agilent 6550 iFunnel time-of-flight MS analyzer. Chromatography of metabolites utilized aqueous normal phase (ANP) chromatography on a Diamond Hydride column (Microsolv). Mobile phases consisted of: (A) 50% isopropanol, containing 0.025% acetic acid, and (B) 90% acetonitrile containing 5 mM ammonium acetate. To eliminate the interference of metal ions on chromatographic peak integrity and electrospray ionization, EDTA was added to the mobile phase at a final concentration of 6 µM. The following gradient was applied: 0-1.0 min, 99% B; 1.0-15.0 min, to 20% B; 15.0 to 29.0, 0% B; 29.1 to 37min, 99% B. Mass spectrometer parameters used for both positive and negative ion mode data acquisition were as follows: drying gas temperature was 200°C with a flow rate of 14 L/min. Nebulizer pressure was at 35 psi. Sheath gas temperature was 350°C with a flow rate of 11 L/min. Capillary and nozzle voltage were at 3500V and 1000V, respectively. Mass spectra were acquired at a rate of 2 spectra/sec over the mass range of 100 to 1700 m/z. Data processing comments: Raw LC/MS data were analyzed using MassHunter Profinder 8.0 and MassProfiler Professional (MPP) 15.1 software (Agilent Technologies). To ascertain the identities of metabolites, LC/MS data were searched against an in-house annotated personal metabolite database created using MassHunter PCDL manager 8.0 (Agilent) based on monoisotopic neutral mass (<5 ppm mass accuracy) and chromatographic retention times of pure standards. A molecular formula generator (MFG) algorithm in MPP was used to generate and score empirical molecular formulae, based on a weighted consideration of monoisotopic mass accuracy, isotope abundance ratios, and spacing between isotope peaks. A tentative compound ID was assigned when the PCDL database and MFG scores concurred for a given candidate molecule. Software procedures used for feature assignments: Tentatively assigned molecules were confirmed based on a match of LC retention times and/or MS/MS fragmentation spectra for pure molecular standards. |
| Ion Mode: | POSITIVE |
| MS ID: | MS004298 |
| Analysis ID: | AN004551 |
| Instrument Name: | Agilent 6550 QTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | MS acquisition comments: For aqueous normal phase LC/MS-based metabolomics, amniotic fluid metabolites were extracted by addition of 1 part amniotic fluid to 15 parts 70% acetonitrile in ddH2O (vol:vol). The mixture was briefly vortexed and then centrifuged for 5 min at 16,000 × g to pellet precipitated proteins. The protein pellet was solubilized in 0.2M NaOH and quantified by DC Protein Assay (Bio-Rad). The volume of metabolite extract was normalized by amniotic protein content. An aliquot of the resulting extract (3 mL) was subjected to LC/MS untargeted metabolite profiling in positive and negative ion modes as described previously described,64 using a platform comprised of an Agilent Model 1290 Infinity II liquid chromatography system coupled to an Agilent 6550 iFunnel time-of-flight MS analyzer. Chromatography of metabolites utilized aqueous normal phase (ANP) chromatography on a Diamond Hydride column (Microsolv). Mobile phases consisted of: (A) 50% isopropanol, containing 0.025% acetic acid, and (B) 90% acetonitrile containing 5 mM ammonium acetate. To eliminate the interference of metal ions on chromatographic peak integrity and electrospray ionization, EDTA was added to the mobile phase at a final concentration of 6 µM. The following gradient was applied: 0-1.0 min, 99% B; 1.0-15.0 min, to 20% B; 15.0 to 29.0, 0% B; 29.1 to 37min, 99% B. Mass spectrometer parameters used for both positive and negative ion mode data acquisition were as follows: drying gas temperature was 200°C with a flow rate of 14 L/min. Nebulizer pressure was at 35 psi. Sheath gas temperature was 350°C with a flow rate of 11 L/min. Capillary and nozzle voltage were at 3500V and 1000V, respectively. Mass spectra were acquired at a rate of 2 spectra/sec over the mass range of 100 to 1700 m/z. Data processing comments: Raw LC/MS data were analyzed using MassHunter Profinder 8.0 and MassProfiler Professional (MPP) 15.1 software (Agilent Technologies). To ascertain the identities of metabolites, LC/MS data were searched against an in-house annotated personal metabolite database created using MassHunter PCDL manager 8.0 (Agilent) based on monoisotopic neutral mass (<5 ppm mass accuracy) and chromatographic retention times of pure standards. A molecular formula generator (MFG) algorithm in MPP was used to generate and score empirical molecular formulae, based on a weighted consideration of monoisotopic mass accuracy, isotope abundance ratios, and spacing between isotope peaks. A tentative compound ID was assigned when the PCDL database and MFG scores concurred for a given candidate molecule. Software procedures used for feature assignments: Tentatively assigned molecules were confirmed based on a match of LC retention times and/or MS/MS fragmentation spectra for pure molecular standards. |
| Ion Mode: | NEGATIVE |
