Summary of study ST001149

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000768. The data can be accessed directly via it's Project DOI: 10.21228/M8DH49 This work is supported by NIH grant, U2C- DK119886.

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Study IDST001149
Study TitlePlasmodium Niemann-Pick Type C1-Related Protein is a Druggable Target Required for Parasite Membrane Homeostasis
Study TypeDrug Exposure
Study SummaryMetabolic examination/confirmation of resistance mutations and GFP tagging observed in study
Institute
Pennsylvania State University
DepartmentBMB
LaboratoryLlinás Laboratory
Last NameLlinás
First NameManuel
AddressW-122 Millennium Science Complex, Pollock Road
Emaileso5005@psu.edu
Phone8148673527
Submit Date2019-02-15
Raw Data AvailableYes
Raw Data File Type(s).xlsx
Analysis Type DetailLC-MS
Release Date2019-05-15
Release Version1
Manuel Llinás Manuel Llinás
https://dx.doi.org/10.21228/M8DH49
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR000768
Project DOI:doi: 10.21228/M8DH49
Project Title:Plasmodium Niemann-Pick Type C1-Related Protein is a Druggable Target Required for Parasite Membrane Homeostasis
Project Type:LC-MS Metabolomics
Project Summary:Hydrophilic LC-MS analysis of PfNCR1 Targeting compounds as part of functional validation of publication
Institute:Pennsylvania State University
Department:BMB
Laboratory:Llinás Laboratory
Last Name:Llinás
First Name:Manuel
Address:W-122 Millennium Science Complex, Pollock Road, University Park, PA 16802
Email:manuel@psu.edu
Phone:Office: (814) 867-3444, Lab: (814) 867-3527
Publications:Plasmodium Niemann-Pick Type C1-Related Protein is a Druggable Target Required for Parasite Membrane Homeostasis
Contributors:Eva S. Istvan, Sudipta Das, Suyash Bhatnagar, Josh R. Beck, Edward Owen, Manuel Llinás, Suresh M. Ganesan, Jacquin C. Niles, Elizabeth A. Winzeler, Akhil B. Vaidya, Daniel E. Goldberg

Subject:

Subject ID:SU001214
Subject Type:Cultured cells
Subject Species:Plasmodium Falciparum
Taxonomy ID:5833

Factors:

Subject type: Cultured cells; Subject species: Plasmodium Falciparum (Factor headings shown in green)

mb_sample_id local_sample_id Drug treatment
SA079821ATQ-20180530Atovaquone, 2.5 hours
SA079822ATQ-MB2Atovaquone, 2.5 hours
SA079823ATQ-MB1Atovaquone, 2.5 hours
SA079824ATQ-20181025Atovaquone, 2.5 hours
SA079825MMV009108-MB2MMV009108, 2.5 hours
SA079826MMV009108-20180530MMV009108, 2.5 hours
SA079827MMV019662-MB1MMV019662, 2.5 hours
SA079828MMV019662-20180530MMV019662, 2.5 hours
SA079829MMV028038-20180530MMV028038, 2.5 hours
SA079830MMV028038-20181025MMV028038, 2.5 hours
SA079831ND-MB2Untreated, 2.5 hours
SA079832ND-20181025Untreated, 2.5 hours
SA079833ND-20180530Untreated, 2.5 hours
SA079834ND-MB1Untreated, 2.5 hours
Showing results 1 to 14 of 14

Collection:

Collection ID:CO001208
Collection Summary:Aspiration of 4 mL media, transfer of sample (1 mL) to 1.5 mL tubes, centrifugation, removal of supernatant, PBS wash, centrifugation, and extraction via 90% methanol with labelled aspartate.
Collection Protocol Filename:Metabolite_Extraction_for_LCMS_2017_Llinas.pdf
Sample Type:Cultured cells
Collection Method:Aspiration of 4 mL media, transfer of sample (1 mL) to 1.5 mL tubes
Collection Location:Biological safety hood
Collection Frequency:once after 2.5 hours treatment
Collection Duration:Collection and extraction occurs in under 20 minutes per plate
Volumeoramount Collected:1 mL containing 1x10^8 infected cells
Storage Conditions:On ice
Collection Vials:1.5 mL eppendorf
Storage Vials:1.5 mL eppendorf

Treatment:

Treatment ID:TR001229
Treatment Summary:Infected Red Blood cells were exposed to Antimalarial Compounds, associated with sample names. All samples were treated with 10xIC50 of a compound for 2.5 hours or left untreated for 2.5 hours.
Treatment Protocol ID:Metabolite_Extraction_for_LCMS_2017_Llinas
Treatment Protocol Filename:Metabolite_Extraction_for_LCMS_2017_Llinas.pdf
Treatment:Abiotic
Treatment Compound:ATQ, MMV009108, MMV019662, MMV028038
Treatment Route:Addition to media
Treatment Dose:10xIC50, 2-10xIC50 for archival data
Treatment Dosevolume:Always equal to or less than 10 uL per 5 mL sample
Treatment Doseduration:2.5 hours
Treatment Vehicle:Stocks kept in either DMSO or Water
Cell Storage:Temperature/Gas Mixture controlled incubator
Cell Growth Container:6-well plate
Cell Growth Config:5 mL per well, each with 1x10^8 cells
Cell Media:RPMI 1640 w/ Albumax, Gentamycin, Hypoxanthine, HEPES, Sodium Bicarbonate
Cell Harvesting:PBS Wash followed by 90% methanol extraction

Sample Preparation:

Sampleprep ID:SP001222
Sampleprep Summary:Samples washed in PBS, spun down, aspirated, extracted in 90% methanol with labelled aspartate, then dried, resuspended in water labelled with chlorpropamide
Sampleprep Protocol Filename:Metabolite_Extraction_for_LCMS_2017_Llinas.pdf
Processing Method:Wash, Lysis, Drying, resuspension
Processing Storage Conditions:On ice
Extraction Method:90% Methanol Lysis
Extract Storage:-80℃
Sample Resuspension:100 uL water with chlorpropamide
Sample Spiking:Labelled Aspartate, Chlorpropamide

Combined analysis:

Analysis ID AN001896
Analysis type MS
Chromatography type Normal phase
Chromatography system Thermo Dionex Ultimate 3000
Column XSelect T3 XP Column Reversed-Phase 2.5 µm Spherical Silica (2.1 mm x 100 mm)
MS Type ESI
MS instrument type Orbitrap
MS instrument name Thermo Exactive Plus Orbitrap
Ion Mode NEGATIVE
Units Peak Area Post-Blank Subtraction

Chromatography:

Chromatography ID:CH001373
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:XSelect T3 XP Column Reversed-Phase 2.5 µm Spherical Silica (2.1 mm x 100 mm)
Chromatography Type:Normal phase

MS:

MS ID:MS001752
Analysis ID:AN001896
Instrument Name:Thermo Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Blank subtraction, RSD cutoffs both by RSD per sample and pooled QCs to eliminate metabolites affected by time-dependency
Ion Mode:NEGATIVE
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