Summary of Study ST000456

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 PR000352. The data can be accessed directly via it's Project DOI: 10.21228/M8JK5M This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

Study ID	ST000456
Study Title	Metabolomic changes during active immunization with anti-METH vaccines.
Study Summary	A targeted metabolomics analysis was performed on serum samples from rats immunized with a methamphetamine-like hapten covalently bound to keyhole limpet hemocyanin (KLH) monomers, and controls. The subjects were approximately 300 g adult male Sprague Dawley rats. RTI International RCMRC received pre-immune serum from RI 11-03A (n=12) which is a matched control serum that was collected before starting immunizations, and week 17 bleeds from RI 11-03A (n=12), RI 11-03B (n=8), and RI 11-03C (n=12). The A-C nomenclature denotes A) complete antigen with adjuvant, B) complete antigen without adjuvant and C) only KLH carrier protein and adjuvant without a METH-like hapten. The rats’ serum samples were extracted and prepared for a combined flow injection (FIA) and LC-MS/MS assay (Biocrates AbsoluteIDQ® p180 kit). The Biocrates AbsoluteIDQ® p180 kit (Biocrates Life Sciences AG, Innsbruck, Austria) can quantitatively measure ~188 biologically relevant metabolites from five analyte groups (acylcarnitines, amino acids, biogenic amines, glycerophospho- and sphingolipids, and hexoses) on a triple quadrupole mass spectrometer.
Institute	University of North Carolina
Department	Discovery-Sciences-Technology (DST)
Laboratory	Sumner Lab
Last Name	Sumner
First Name	Susan
Address	Eastern Regional Comprehensive Metabolomics Resource Core, UNC Nutrition Research Institute, 500 Laureate Way, Kannapolis, NC, 28081
Email	susan_sumner @unc.edu
Phone	704-250-5066
Submit Date	2016-08-31
Num Groups	4
Total Subjects	44
Raw Data Available	Yes
Raw Data File Type(s)	wiff
Analysis Type Detail	LC-MS
Release Date	2017-10-03
Release Version	1

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Project:

Project ID:	PR000352
Project DOI:	doi: 10.21228/M8JK5M
Project Title:	Metabolomic changes during active immunization with anti-METH vaccines.
Project Summary:	Metabolomics studies will be conducted on serum samples from rats immunized with a methamphetamine conjugate vaccine. The Subjects were approximately 300 g adult male Sprague Dawley rats. The following samples were sent to RTI International RCMRC: Pre-immune serum from RI 11-03A (n=12) which is a matched control serum that was collected before starting immunizations, and week 17 bleeds from RI 11-03A (n=12), RI 11-03B (n=8), and RI 11-03C (n=12). The A-C nomenclature denotes A) complete antigen with adjuvant, B) complete antigen without adjuvant and C) only KLH carrier protein and adjuvant without a METH-like hapten.
Institute:	University of Arkansas
Department:	Department of Pharmacology and Toxicology
Last Name:	Owens
First Name:	S. Michael
Address:	4301 W. Markham St., #611 Little Rock, AR 72205 USA
Email:	mowens@uams.edu
Phone:	501-686-5487

Subject:

Subject ID:	SU000477
Subject Type:	Animal
Subject Species:	Rattus norvegicus
Taxonomy ID:	10116
Species Group:	Mammal

Factors:

Subject type: Animal; Subject species: Rattus norvegicus (Factor headings shown in green)

mb_sample_id	local_sample_id	MCV	Adjuvant	Sample Collection
SA023136	MO_38	Act KLH	RIBI	Immunized (17 wks)
SA023137	MO_44	Act KLH	RIBI	Immunized (17 wks)
SA023138	MO_41	Act KLH	RIBI	Immunized (17 wks)
SA023139	MO_42	Act KLH	RIBI	Immunized (17 wks)
SA023140	MO_34	Act KLH	RIBI	Immunized (17 wks)
SA023141	MO_43	Act KLH	RIBI	Immunized (17 wks)
SA023142	MO_33	Act KLH	RIBI	Immunized (17 wks)
SA023143	MO_39	Act KLH	RIBI	Immunized (17 wks)
SA023144	MO_40	Act KLH	RIBI	Immunized (17 wks)
SA023145	MO_36	Act KLH	RIBI	Immunized (17 wks)
SA023146	MO_35	Act KLH	RIBI	Immunized (17 wks)
SA023147	MO_37	Act KLH	RIBI	Immunized (17 wks)
SA023112	P17A_2	-	-	-
SA023113	PBS_03	-	-	-
SA023114	p180 Cal7	-	-	-
SA023115	p180 Cal6	-	-	-
SA023116	p180-MetaDis QC1	-	-	-
SA023117	P0_1	-	-	-
SA023118	P17C_1	-	-	-
SA023119	P17B_1	-	-	-
SA023120	p180 Cal4	-	-	-
SA023121	p180 Cal3	-	-	-
SA023122	TP_1	-	-	-
SA023123	p180 Cal5	-	-	-
SA023124	p180 Cal2	-	-	-
SA023125	p180 Cal1	-	-	-
SA023126	p180-MetaDis QC3	-	-	-
SA023127	P17A_1	-	-	-
SA023128	PBS_01	-	-	-
SA023129	p180-MetaDis QC2	-	-	-
SA023130	TP_2	-	-	-
SA023131	P17B_2	-	-	-
SA023132	TP_3	-	-	-
SA023133	P17C_2	-	-	-
SA023134	PBS_02	-	-	-
SA023135	P0_2	-	-	-
SA023148	MO_28	SSM09	None	Immunized (17 wks)
SA023149	MO_26	SSM09	None	Immunized (17 wks)
SA023150	MO_32	SSM09	None	Immunized (17 wks)
SA023151	MO_29	SSM09	None	Immunized (17 wks)
SA023152	MO_27	SSM09	None	Immunized (17 wks)
SA023153	MO_25	SSM09	None	Immunized (17 wks)
SA023154	MO_31	SSM09	None	Immunized (17 wks)
SA023155	MO_30	SSM09	None	Immunized (17 wks)
SA023156	MO_21	SSM09	RIBI	Immunized (17 wks)
SA023157	MO_17	SSM09	RIBI	Immunized (17 wks)
SA023158	MO_20	SSM09	RIBI	Immunized (17 wks)
SA023159	MO_16	SSM09	RIBI	Immunized (17 wks)
SA023160	MO_14	SSM09	RIBI	Immunized (17 wks)
SA023161	MO_19	SSM09	RIBI	Immunized (17 wks)
SA023162	MO_24	SSM09	RIBI	Immunized (17 wks)
SA023163	MO_22	SSM09	RIBI	Immunized (17 wks)
SA023164	MO_18	SSM09	RIBI	Immunized (17 wks)
SA023165	MO_13	SSM09	RIBI	Immunized (17 wks)
SA023166	MO_23	SSM09	RIBI	Immunized (17 wks)
SA023167	MO_15	SSM09	RIBI	Immunized (17 wks)
SA023168	MO_1	SSM09	RIBI	Pre-Immune (0 wks)
SA023169	MO_10	SSM09	RIBI	Pre-Immune (0 wks)
SA023170	MO_8	SSM09	RIBI	Pre-Immune (0 wks)
SA023171	MO_12	SSM09	RIBI	Pre-Immune (0 wks)
SA023172	MO_11	SSM09	RIBI	Pre-Immune (0 wks)
SA023173	MO_9	SSM09	RIBI	Pre-Immune (0 wks)
SA023174	MO_5	SSM09	RIBI	Pre-Immune (0 wks)
SA023175	MO_2	SSM09	RIBI	Pre-Immune (0 wks)
SA023176	MO_3	SSM09	RIBI	Pre-Immune (0 wks)
SA023177	MO_7	SSM09	RIBI	Pre-Immune (0 wks)
SA023178	MO_4	SSM09	RIBI	Pre-Immune (0 wks)
SA023179	MO_6	SSM09	RIBI	Pre-Immune (0 wks)

Showing results 1 to 68 of 68

Showing results 1 to 68 of 68

Collection:

Collection ID:	CO000471
Collection Summary:	Serum Samples from Rats Immunized with a Methamphetamine Conjugate Vaccine
Sample Type:	Blood

Treatment:

Treatment ID:	TR000491
Treatment Summary:	Treatment A - complete antigen with adjuvant

Sample Preparation:

Sampleprep ID:	SP000484
Sampleprep Summary:	Study Phenotypic Pools Thawed serum samples were vortexed and centrifuged at 5,000 rcf for 4 minutes. The following phenotypic pools were created by combining 10 µL from each phenotypic sample into a 2 mL LoBind eppendorf tube: 1. Pre-immune serum from RI 11-03A (n=12) which is a matched control serum that was collected before starting immunizations. The final volume of this phenotypic pool was 120 µL which was vortexed for 30 seconds. Two phenotypic pools (0wk-03A_Pool1 and 0wk-03A_Pool2) of 25 µL each were created. 2. Week 17 bleeds from RI 11-03A (n=12). The final volume of this phenotypic pool was 120 µL which was vortexed for 30 seconds. Two phenotypic pools (17wk-03A_Pool1 and 17wk-03A_Pool1) of 25 µL each were created. 3. RI 11-03B (n=8).The final volume of this phenotypic pool was 80 µL which was vortexed for 30 seconds. Two phenotypic pools (17wk-03B_Pool1 and 17wk-03B_Pool2) of 25 µL each were created. 4. and RI 11-03C (n=12). The final volume of this phenotypic pool was 120 µL which was vortexed for 30 seconds. Two phenotypic pools (17wk-03C_Pool1and 17wk-03C_Pool2) of 25 µL each were created. Whole Study Pooled QC Whole study pooled QC samples were prepared by combining 25 µL of each phenotypic pool sample created above into a 2 mL LoBind eppendorf tube. This QC pooled sample was then vortexed for 30 sec. Then, three whole study pooled QC samples of 25 µL each were aliquoted into 2 mL LoBind eppendorf tubes. Biocrates Plate Preparation: All samples (Phenotypic pools, whole study pooled QC and study samples) were vortexed for 10 minutes at 5,000 rpm. Samples were then centrifuged at 16,000 rcf for 4 minutes and at 4 °C. A Biocrates p180 kit was prepared following the AbsoluteIDQ™ p180 Kit metabolomics procedure. Briefly, an internal standard mix was added to 95 of the 96 wells. Next, zero samples, QC standards and calibration standards were added to their corresponding wells. The study samples, phenotypic and whole study pooled QC Samples (20 µL) were then added to the appropriate wells and dried for 30 minutes under nitrogen flow. The plate was derivatized using a 5% phenylisothiocyanate (PITC) solution in (1:1:1) ethanol:pyridine:water (v/v/v) and, then, incubated for 20 minutes at room temperature followed by a drying step under nitrogen flow. An extraction solvent (5 mM ammonium acetate in methanol) was added to all wells. The plate was then shaken and centrifuged. After centrifugation, 150 µL was removed and transferred to a second 96-well plate (LCMS plate). This second plate was diluted with 150 µL of HPLC grade water for a subsequent LCMS (MRM analysis) for measuring amino acids and biogenic amines. All wells in the original plate were diluted with 400 µL of flow injection analysis (FIA) Running Solvent for a FIA-MS (MRM analysis) for measuring lipids, acylcarnitines, and hexose.

Sampleprep ID:

SP000484

Sampleprep Summary:

Study Phenotypic Pools Thawed serum samples were vortexed and centrifuged at 5,000 rcf for 4 minutes. The following phenotypic pools were created by combining 10 µL from each phenotypic sample into a 2 mL LoBind eppendorf tube: 1. Pre-immune serum from RI 11-03A (n=12) which is a matched control serum that was collected before starting immunizations. The final volume of this phenotypic pool was 120 µL which was vortexed for 30 seconds. Two phenotypic pools (0wk-03A_Pool1 and 0wk-03A_Pool2) of 25 µL each were created. 2. Week 17 bleeds from RI 11-03A (n=12). The final volume of this phenotypic pool was 120 µL which was vortexed for 30 seconds. Two phenotypic pools (17wk-03A_Pool1 and 17wk-03A_Pool1) of 25 µL each were created. 3. RI 11-03B (n=8).The final volume of this phenotypic pool was 80 µL which was vortexed for 30 seconds. Two phenotypic pools (17wk-03B_Pool1 and 17wk-03B_Pool2) of 25 µL each were created. 4. and RI 11-03C (n=12). The final volume of this phenotypic pool was 120 µL which was vortexed for 30 seconds. Two phenotypic pools (17wk-03C_Pool1and 17wk-03C_Pool2) of 25 µL each were created. Whole Study Pooled QC Whole study pooled QC samples were prepared by combining 25 µL of each phenotypic pool sample created above into a 2 mL LoBind eppendorf tube. This QC pooled sample was then vortexed for 30 sec. Then, three whole study pooled QC samples of 25 µL each were aliquoted into 2 mL LoBind eppendorf tubes. Biocrates Plate Preparation: All samples (Phenotypic pools, whole study pooled QC and study samples) were vortexed for 10 minutes at 5,000 rpm. Samples were then centrifuged at 16,000 rcf for 4 minutes and at 4 °C. A Biocrates p180 kit was prepared following the AbsoluteIDQ™ p180 Kit metabolomics procedure. Briefly, an internal standard mix was added to 95 of the 96 wells. Next, zero samples, QC standards and calibration standards were added to their corresponding wells. The study samples, phenotypic and whole study pooled QC Samples (20 µL) were then added to the appropriate wells and dried for 30 minutes under nitrogen flow. The plate was derivatized using a 5% phenylisothiocyanate (PITC) solution in (1:1:1) ethanol:pyridine:water (v/v/v) and, then, incubated for 20 minutes at room temperature followed by a drying step under nitrogen flow. An extraction solvent (5 mM ammonium acetate in methanol) was added to all wells. The plate was then shaken and centrifuged. After centrifugation, 150 µL was removed and transferred to a second 96-well plate (LCMS plate). This second plate was diluted with 150 µL of HPLC grade water for a subsequent LCMS (MRM analysis) for measuring amino acids and biogenic amines. All wells in the original plate were diluted with 400 µL of flow injection analysis (FIA) Running Solvent for a FIA-MS (MRM analysis) for measuring lipids, acylcarnitines, and hexose.

Combined analysis:

Analysis ID	AN000713	AN000714	AN000715
Analysis type	MS	MS	MS
Chromatography type	Reversed phase	Reversed phase	Reversed phase
Chromatography system	Agilent 1100	Agilent 1100	Agilent 1100
Column	Agilent Eclipse XDB-C18 (100 x 3.0mm,3.5um)	Agilent Eclipse XDB-C18 (100 x 3.0mm,3.5um)	Agilent Eclipse XDB-C18 (100 x 3.0mm,3.5um)
MS Type	ESI	ESI	ESI
MS instrument type	Triple quadrupole	Triple quadrupole	Triple quadrupole
MS instrument name	ABI Sciex API 4000 QTrap	ABI Sciex API 4000 QTrap	ABI Sciex API 4000 QTrap
Ion Mode	POSITIVE	POSITIVE	NEGATIVE
Units	Intensity	Intensity	Intensity

Chromatography:

Chromatography ID:	CH000514
Instrument Name:	Agilent 1100
Column Name:	Agilent Eclipse XDB-C18 (100 x 3.0mm,3.5um)
Column Temperature:	50 degrees C
Flow Rate:	0.5mL/min
Chromatography Type:	Reversed phase

MS:

MS ID:	MS000633
Analysis ID:	AN000713
Instrument Name:	ABI Sciex API 4000 QTrap
Instrument Type:	Triple quadrupole
MS Type:	ESI
Ion Mode:	POSITIVE

MS ID:	MS000634
Analysis ID:	AN000714
Instrument Name:	ABI Sciex API 4000 QTrap
Instrument Type:	Triple quadrupole
MS Type:	ESI
Ion Mode:	POSITIVE

MS ID:	MS000635
Analysis ID:	AN000715
Instrument Name:	ABI Sciex API 4000 QTrap
Instrument Type:	Triple quadrupole
MS Type:	ESI
Ion Mode:	NEGATIVE