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MB Sample ID: SA098168

Local Sample ID:CKD014
Subject ID:SU001427
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57BL6J
Age Or Age Range:18-20 weeks
Weight Or Weight Range:20-30g
Gender:Male and female
Animal Animal Supplier:Jackson Laboratories
Animal Housing:5/cage
Animal Light Cycle:12h
Animal Feed:Ad libitum. Control mice received custom casein-diet. Chronic kidney disease was induced by supplementing casein-based diet with 0.15% adenine
Animal Water:Ad libitum

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Subject:

Subject ID:SU001427
Subject Type:Mammal
Subject Species:Mus musculus
Taxonomy ID:10090
Genotype Strain:C57BL6J
Age Or Age Range:18-20 weeks
Weight Or Weight Range:20-30g
Gender:Male and female
Animal Animal Supplier:Jackson Laboratories
Animal Housing:5/cage
Animal Light Cycle:12h
Animal Feed:Ad libitum. Control mice received custom casein-diet. Chronic kidney disease was induced by supplementing casein-based diet with 0.15% adenine
Animal Water:Ad libitum

Factors:

Local Sample IDMB Sample IDFactor Level IDLevel ValueFactor Name
CKD014SA098168FL013863CKDGroup

Collection:

Collection ID:CO001422
Collection Summary:Skeletal muscle was quickly dissected, trimmed of fat and connective tissues and rinsed in PBS to remove and blood. Dissection occurred under ketamine/xylazine anesthesia and snap frozen in liquid nitrogen. Frozen muscles were stored at -80C until processing.
Sample Type:Muscle
Storage Conditions:-80℃

Treatment:

Treatment ID:TR001442
Treatment Summary:We utilized an established adenine-diet model to induce CKD in mice. Mice were assigned to a casein-based chow diet for 7 days, followed by induction of renal tubular injury by supplementing the diet with 0.2% adenine for 7 days, and were subsequently maintained on a 0.15% adenine diet for 7 more weeks. CKD mice were then placed back on control casein diet for 2 weeks to prior to euthanasia and terminal experiments. Control mice received casein diet for the duration of the study.
Animal Anesthesia:Ketamine/Xylazine
Animal Endp Euthanasia:Ketamine/Xylazine

Sample Preparation:

Sampleprep ID:SP001435
Sampleprep Summary:Muscles were thawed on ice, weighed, and homogenized with a Teflon-tipped conical pestle with a metal rod (Micro-Tube Sample Pestle with Conical Teflon Tip, fits 1.5ml Tubes, autoclavable at 121°F; Research Products International Corp; 199221; Fisher Scientific). The pestle was rinsed with 2-Propanol, water, and methanol and patted dry with a KimWipe in between samples. The samples were centrifuged to pellet the tissue debris and protein concentrations were quantified on the QuBit. The samples were normalized to 500µg/mL of protein with 5mM Ammonium Acetate in water prior to extraction for a total volume of 100µL. 25µL of sample was aliquoted into a clean tube and extracted with 5µL of Global Metabolomics IS and 200µL of 8:1:1 Acetonitrile:Methanol:Acetone to precipitate proteins. The samples were incubated at 4°C for 30 min and centrifuged at 20,000xg at 4°C for 10min. 200µL of supernatant was transferred to a clean Eppendorf tube and dried under nitrogen gas at 30°C and then reconstituted at 25µL in Global Metabolomics Inj. Std. Mix.

Combined analysis:

Analysis ID AN002251 AN002252
Analysis type MS MS
Chromatography type Reversed phase Reversed phase
Chromatography system Thermo Dionex Thermo Dionex
Column ACE 5 C18-300 (100 x 2.1mm) ACE 5 C18-300 (100 x 2.1mm)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Exactive Plus Orbitrap
Ion Mode POSITIVE NEGATIVE
Units peak height peak height

Chromatography:

Chromatography ID:CH001655
Chromatography Summary:samples were processed on a Thermo Q-Exactive Oribtrap mass spectrometer with Dionex UHPLC and autosampler. All samples were analyzed in positive and negative heated electrospray ionization with a mass resolution of 35,000 at m/z 200 as separate injections. Separation was achieved on an ACE 18-pfp 100 x 2.1 mm, 2 µm column with mobile phase A as 0.1% formic acid in water and mobile phase B as acetonitrile. This is a polar embedded stationary phase that provides comprehensive coverage, but does have some limitation is the coverage of very polar species. The flow rate was 350 µL/min with a column temperature of 25°C. 4 µL was injected for negative ions and 2 µL for positive ions.
Instrument Name:Thermo Dionex
Column Name:ACE 5 C18-300 (100 x 2.1mm)
Column Temperature:25C
Flow Rate:350ul/min
Sample Injection:4ul for negative ion, 2ul for positive ion
Solvent A:100% water; 0.1% formic acid
Solvent B:100% acetonitrile
Chromatography Type:Reversed phase

MS:

MS ID:MS002096
Analysis ID:AN002251
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:samples were processed on a Thermo Q-Exactive Oribtrap mass spectrometer with Dionex UHPLC and autosampler. All samples were analyzed in positive and negative heated electrospray ionization with a mass resolution of 35,000 at m/z 200 as separate injections. Separation was achieved on an ACE 18-pfp 100 x 2.1 mm, 2 µm column with mobile phase A as 0.1% formic acid in water and mobile phase B as acetonitrile. This is a polar embedded stationary phase that provides comprehensive coverage, but does have some limitation is the coverage of very polar species. The flow rate was 350 µL/min with a column temperature of 25°C. 4 µL was injected for negative ions and 2 µL for positive ions. Data from positive and negative ion modes were separately subjected to statistical analyses. MZmine (freeware) was used to identify features, deisotope, align features and perform gap filling to fill in any features that may have been missed in the first alignment algorithm. All adducts and complexes were identified and removed from the data set. The primary source of feature identification was performed by mapping against an internal retention time metabolite library established by the SECIM. Additional metabolite searches were performed using HMDB (http://www.hmdb.ca) and the Metabolomics Workbench (https://www.metabolomicsworkbench.org) through a search of the m/z ratio with a [M+H] adduct and a tolerance of 0.002 m/z.
Ion Mode:POSITIVE
  
MS ID:MS002097
Analysis ID:AN002252
Instrument Name:Thermo Exactive Plus Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:samples were processed on a Thermo Q-Exactive Oribtrap mass spectrometer with Dionex UHPLC and autosampler. All samples were analyzed in positive and negative heated electrospray ionization with a mass resolution of 35,000 at m/z 200 as separate injections. Separation was achieved on an ACE 18-pfp 100 x 2.1 mm, 2 µm column with mobile phase A as 0.1% formic acid in water and mobile phase B as acetonitrile. This is a polar embedded stationary phase that provides comprehensive coverage, but does have some limitation is the coverage of very polar species. The flow rate was 350 µL/min with a column temperature of 25°C. 4 µL was injected for negative ions and 2 µL for positive ions. Data from positive and negative ion modes were separately subjected to statistical analyses. MZmine (freeware) was used to identify features, deisotope, align features and perform gap filling to fill in any features that may have been missed in the first alignment algorithm. All adducts and complexes were identified and removed from the data set. The primary source of feature identification was performed by mapping against an internal retention time metabolite library established by the SECIM. Additional metabolite searches were performed using HMDB (http://www.hmdb.ca) and the Metabolomics Workbench (https://www.metabolomicsworkbench.org) through a search of the m/z ratio with a [M+H] adduct and a tolerance of 0.002 m/z.
Ion Mode:NEGATIVE
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