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Providing cutting edge biochemistry methods and molecular biology techniques for nutrition research

Harvard Medical School

Service Description

Metabolic Phenotyping Core

Providing users with access to high-quality model creation and detailed metabolic phenotyping. Services include:

  • Immortalized cells, primary adipocytes or hepatocytes subjected to
  • Compound, drug, or metabolite-treated
  • siRNA or shRNA-mediated gene knockdown
  • CRISPR-mediated gene knockout
  • CRISPR-mediated sequence specific alteration,
  • Overexpression of genes of interest
  • Seahorse mitochondrial assays
  • Biochemical insulin signaling assay
  • Hepatic glucose output and lipolysis assays

Have Questions? Contact Us.

Alexander Soukas, MD, PhD, Core Director • Email 

Robert Gerszten, Co-Director • Email

New York

Molecular Biology and Molecular Genetics Core

Service Description

Applying the tools and technologies of molecular genetics and genomics to elucidate the molecular-genetic bases of obesity and its comorbidities.

  • Plasmid Generation (cloning from cDNA, site directed, mutagenesis/plasmid construction)
  • qPCR (reverse transcription, primer design/ordering, qPCR run in duplicate, qPCR machine use)
  • Genotyping (design of restriction digest PCR primers for genotyping, restriction enzyme based genotyping, PCR based genotyping)
  • Microscopy (confocal, fluorescent, neuronal cell counting software)
  • Facs Sorting (design and analysis)

Have Questions? Contact Us.

Wendy Chung, MD, PhD, Director
Email • Phone: (212) 851-5313

University of Colorado Anschutz Medical Campus

Service Description

Molecular and Cellular Analytical Core

Consultation and services for gene expression analysis to investigate molecular targets (metabolic, inflammatory, lipogenic, etc.) influencing nutrition, obesity and maternal-fetal metabolism.

  • RNA isolation from various tissues (liver, muscle, adipose) and cell types (macrophages, cell cultures) from human, nonhuman primate and rodents
  • cDNA synthesis
  • PCR primer design & ordering (we currently have over 600 primer sets for human, mouse and rat genes)
  • Quantitative real-time PCR on genes of interest and two or more reference genes

Have Questions? Contact Us.

Jacob (Jed) E. Friedman, PhD, Core Director
Email • (303) 724-3983

Rachel C. Janssen, MS, Core Manager
Email • (303) 724-3979

Service Description

Molecular and Cellular Analytical Core

Seahorse Bioscience XF24 Extracellular Flux Analyzer simultaneously interrogates two major energy producing pathways of the cell – mitochondrial respiration and glycolysis – in a microplate, in real-time. The XF24-3 Analyzer determines in vitro oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) to assess cellular functions such as oxidative phosphorylation, glycolysis, and fatty acid oxidation. The XF24-3 technology provides the tools to rapidly detect cellular responses to substrates, inhibitors and other compounds in a 24-well-plate format.

Initial training on the Seahorse machine and software is provided and users are expected to determine their experiment design and interpret their own data. Users will supply their own FluxPaks (plates & cartridges), calibrant, reagents and medium. There is a per assay cost for use of the bioanalyzer.

Have Questions? Contact Us.

Jacob (Jed) E. Friedman, PhD, Core Director
Email • (303) 724-3983

Rachel C. Janssen, MS, Core Manager
Email • (303) 724-3979

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Service Description

Molecular and Cellular Analytical Core

Consultation and services for a broad range of assays over a variety of tissue types, fluids and cells for high-throughput study of nutrition and obesity-related disorders.

  • Cytokine analysis by multiplex assays and cytokine arrays from multiple sample types (e.g. breastmilk, serum) in humans, rodents and cell cultures
  • Hormone analysis by ELISA assays for leptin, adiponectin, insulin in human and rodent samples
  • Endotoxin assays to measure circulating endotoxin levels in serum
  • Western blot analysis of targeted and phosphorylated proteins
  • Colorimetric assays for quantitation of proteins, glucose, triglycerides and free fatty acids
  • Mitochondrial activity assays to investigate activity of respiratory chain enzyme complexes I, II, II+III, III, IV, citrate synthase, aconitase and superoxide dismutase.

Have Questions? Contact Us.

Jacob (Jed) E. Friedman, PhD, Core Director
Email • (303) 724-3983

Rachel C. Janssen, MS, Core Manager
Email • (303) 724-3979

University of Michigan

Service Description

Molecular Phenotyping Core

The Seahorse XF instrument is used to assess extracellular O2, pH, glucose and lactate fluxes to provide estimates of glucose utilization in cells and tissues.

Have Questions? Contact Us.

Sydney Bridges, Lab Manager
Email • Phone: (734) 764-2682

University of North Carolina at Chapel Hill

Service Description

Metabolic Molecular Phenotyping Core

IMG_6840Assays for Aqueous Choline Metabolites include choline, betaine, (phosphocholine and glycerophosphocholine) in plasma, serum, urine, breast milk, liquid, food, and tissues. Methionine, and creatinine can also be included, by request, at no extra charge.  Please note that phosphocholine and glycerophosphocholine are too low to measure in plasma.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

Service Description

Metabolic Molecular Phenotyping Core

Analysis of arsenic (metals and metalloids) in biological or environmental samples. Atomic Absorption Spectrometry includes:

  • Oxidation State-Specific Analysis of Arsenic (Aqueous Solutions or Simple Biological Matrices)
    Trivalent and pentavalent arsenic species in aqueous solutions or simple biological matrices (e.g., cell culture medium or urine) are analyzed directly (without extraction or chemical digestion) using hydride generation-cryotrapping-atomic absorption spectrometry (HG-CT-AAS). This method detects and quantifies 7 arsenic species, including arsenite, arsenate, methylarsenite, methylarsenate, dimethylarsenite, dimethylarsenate, and trimethylarsine oxide.
  • Oxidation State-Specific Speciation Analysis of Arsenic (Aqueous Solutions or Simple Biological Matrices) – Low Arsenic Content
    Trivalent and pentavalent arsenic species in aqueous solutions or simple biological matrices (e.g., cell culture medium or urine) are analyzed directly (without extraction or chemical digestion) using HG-CT-inductively coupled with the plasma-mass spectrometer (ICP-MS). This method can detect and quantify very low levels of 7 arsenic species, including arsenite, arsenate, methylarsenite, methylarsenate, dimethylarsenite, dimethylarsenate, and trimethylarsine oxide.
  • Analysis of Total Arsenic or Selenium (Aqueous Solutions or Simple Biological Matrices)
    Total arsenic or selenium in aqueous solutions or simple biological matrices (e.g., cell culture medium or urine) is analyzed directly (without extraction or chemical digestion) using HG-CT-AAS or graphite furnace (GF)-AAS, providing only information about the total amount of the element.
  • Analysis of Total Arsenic or Selenium (Complex Biological Matrices)
    Total arsenic or selenium in complex biological matrices (e.g., sediments, cells or tissues) is analyzed after extraction or chemical digestion of samples, using HG-CT-AAS or graphite furnace (GF)-AAS and providing only information about the total amount of the element.
  • Speciation Analysis of Arsenic (Complex Biological Matrices)
    Arsenic species in complex biological matrices (e.g., sediments, cells or tissues) are analyzed after extraction or chemical digestion of samples using HG-CT-AAS. This method detects and quantifies inorganic arsenic, methylarsenic, dimethylarsenic and trimethylarsenic without providing information on the oxidation state of arsenic in the above species.
  • Speciation Analysis of Arsenic (Aqueous Solutions or Simple Biological Matrices)
    Arsenic species in aqueous solutions or simple biological matrices (e.g., cell culture medium or urine) are analyzed directly (without extraction or chemical digestion) using HG-CT-AAS. This method detects and quantifies inorganic arsenic, methylarsenic, dimethylarsenic and trimethylarsenic without providing information on the oxidation state of arsenic in the above species.
  • Speciation Analysis of Arsenic (Aqueous Solutions or Simple Biological Matrices) – Low Arsenic Content
    Arsenic species in aqueous solutions or simple biological matrices (e.g., cell culture medium or urine) are analyzed directly (without extraction or chemical digestion) using HG-CT-inductively coupled with the plasma-mass spectrometer (ICP-MS). This method can detect and quantify very low levels of inorganic arsenic, methylarsenic, dimethylarsenic and trimethylarsenic without providing information on the oxidation state of arsenic in the above species.

Have Questions? Contact Us.

Mirek Styblo, PhD, Director, Atomic Absorption Spectroscopy Lab
Email • Phone: (919) 966-5721

 

 

Service Description

Metabolic Molecular Phenotyping Core

Assays for Choline Metabolites (choline, betaine, phosphatidylcholine, sphingomyelinin, phosphocholine, and glycerophosphocholine) in plasma, serum, urine, breast milk food and tissues. Methionine, and creatinine can also be included, by request, at no extra charge. Please note that phosphocholine and glycerophosphocholine are too low to measure in plasma.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

The Dimethyl-glycine, Choline and Betaine assay is designed to measure the metabolites listed above by liquid chromatography-stable isotope dilution-multiple reaction monitoring mass spectrometry (LC-SID-MRM/MS). Plasma, urine, food and/or tissue samples may be submitted.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

Service Description

Metabolic Molecular Phenotyping Core

This is an hourly rate for use of the Seahorse XF24 Platform. This assumes prior training or experience with the XF24 Analyzer and associated software. Users are expected to supply their own plates, reagents, calibrant and medium. All experiments are run at the Nutrition Research Institute in Kannapolis, NC.

Seahorse sells kits for assessing glycolytic function (Cellular Glycolysis Test) and mitochondrial function (Mitochondrial Function Test). For accurate results, the user should plan to spend one day assessing cell density, a second day to titrate kit reagents and your test reagents, and a third day (or more) for actual experiments.

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

 

Service Description

Metabolic Molecular Phenotyping Core

Drug Abuse Tests includes analysis for the amount of stimulants, entactogens, hallucinogens, and other drugs (including amphetamines, barbiturates, benzodiazepines, cannabinoids, and cocaine metabolites) that can be found in the human body.

Have Questions? Contact Us.

Qing Shi, Lab Manager-Chapel Hill Facilities
Email • Phone:(919) 966-7162

 

Service Description

Metabolic Molecular Phenotyping Core

FACS Aria III Training typically consists of a 2-hour classroom session and then at least two 5-hour sessions on the instrument. The fee is per person. The user will learn some theory around flow cytometry and cell sorting, and also how to run the Aria III FACS (BD Biosciences). The user is expected to do a lot of reading on their own between the classroom and instruments sessions. Due to limited time, training touches upon but does not focus on the numerous techniques used in flow cytometry such as cell cycle analysis, immunophenotyping, redox state, etc. The user interested in a particular technique will have to research the best strategies for their system, and we will provide guidance.

The Aria III is equipped with blue (488 nm), green (561 nm), red (633 nm) and violet (405 nm) lasers with 11 detectors that can measure up to 10 colors simultaneously. We are currently unable to run un-fixed human or primate cells as the Aria III instrument is not in a BSL2+ facility.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

The Fatty Acid assay is designed to measure fatty acids by thin layer chromatography/gas chromatography (TLC/GC). Plasma, breast milk, food or tissue samples may be submitted. The following  fatty acids can be measured by GC-FID:

FA Name Common Name
C14:0 Myristic acid
C14:1 Myristoleic acid
C16:0 Palmitic acid
C16:1 Palmitoleic acid
C18:0 Stearic acid
C18:1 Oleic acid
C18:2 Linoleic acid
C18:3 Alpha-Linolenic acid
C20:0 Arachidic acid
C20:1 Eicosenoic acid
C20:2 Eicosadienoic acid
C20:3 8,11,14-Eicosatrienoic acid
C20:3 11,14,17-Eicosatrienoic acid
C20:4 Arachidonic acid
C20:5 Eicosapentaenoic acid
C22:0 Behenic acid
C22:1 Erucic acid
C22:5 Docosapentaenoic acid
C22:6 Docosahexaenoic acid
C24:0 Tetracosanoic acid
C24:1 Nervonic acid

 

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

This 96-well format microbiological assay is considered to be one of the most effective methods for measuring folate levels in food or tissue.

This assay measures the growth response of the bacteria L. casei (ATCC 7469) to folic acid. Since these bacteria require folate to grow, the growth response is directly related to the folate level in the food or tissue sample. L. casei bacteria are used because they respond to the widest variety of folate derivatives. 5-formyltetrahydrofolate (5-formyl THF) is the major folic acid derivative found in food products and is therefore used as the standard in this assay. For determination of food folate, a tri-enzyme treatment is necessary to extract the folate in the sample prior to the assay. Protease and α-amylase are used to extract folate bound to protein or carbohydrate matrices. Rat serum conjugase is used to hydrolyzed polyglutamyl forms of folate to monoglutamyl forms which are measured in the microbiological assay. It has been reported that the tri-enzyme treatment consistently yields higher folate values when compared to the more traditional methods, which use either one enzyme or none at all. The coefficient of variation for this assay is 3.9%. The assay yields a linear response between 0 and 120pg.

Red Blood Cells or Plasma

This 96-well format microbiological assay is considered to be one of the most effective methods for measuring folate levels in plasma, serum or red blood cells.

This assay measures the growth response of the bacteria L. casei (ATCC 7469) to folic acid. Since these bacteria require folate to grow, the growth response is directly related to the folate level in the blood or RBC sample. L. casei bacteria are used because they respond to the widest variety of folate derivatives. 5-methyltetrahydrofolate (5-methyl THF) is the major folic acid derivative found in plasma, serum, and red blood cells and is therefore used as the standard in the assay for biological samples. The coefficient of variation for this assay is 3.9%. The assay yields a linear response between 0 and 120pg.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site or folate (food or tissue) or folate (for red blood cells or plasma).

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

Fluorescence Activated Cell Sorting is done on an Aria III (BD Biosciences) fluorescence activated cell sorter (FACS). Sorts can be optimized for purity, yield or single cell modes. The Aria III is equipped with blue (488 nm), green (561 nm), red (633 nm) and violet (405 nm) lasers with 11 detectors that can measure up to 10 colors simultaneously. We are currently unable to sort human or primate cells as the Aria III instrument is not in a BSL2+ facility.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

The Luminex Multiplex Plasma/Serum Assay is a bead-based, multiplex assay system utilizing Luminex® technology and the FlexMap 3D instrument. Multiple analytes in 25 µL (typically) volume of either plasma or serum can be measured in single wells containing the corresponding bead types. Customers are responsible for ordering their own plates.

Multiple analytes in 25 µL (typically) plasma or serum can be measured together in single wells containing the corresponding bead types. While plate costs increase with the number of analytes chosen, multiplexing lowers your overall material and labor costs to perform multiple assays.

The panels listed below are designed by EMD Millipore, although other companies also design assays for the Luminex platform. You can choose one or all of the analytes within a panel to multiplex together. Each 96-well plate will typically handle 38 samples in duplicate. Please visit EMD Millipore to design and price your plates. Please be sure to select only assays that have ‘MAG’ in the title (these are magnetic bead based assays).

You can choose from many kinds of panels including: metabolic hormones, gut hormones, pituitary hormones, bone, CVD, circulating cancer biomarkers, neurodegenerative disease, and neurological disorders. Most assays are for human proteins but many are available for mouse and rat as well.

Please note that the following assays are performed at the Chapel Hill facilities and would not be done at the Nutrition Research Institute:

  • Oxidative Stress includes Catalase, Glutathione Peroxidase (GPx), Glutathione Reductase (GR), Glutathione S-Transferases (GST), Superoxide Dismutase, Thiobarbituric Acid Substances (TBARS), Oxidized and Reduced Glutathione (GSH/GSSG) and Thioredoxin Reductase.
  • Adipokines and Cytokines includes Adiponectin, Interleukin-1 & Interleukin-1 HS, Interleukin-2, Interleukin-6 Interleukin-6 HS and TNF-α & TNF-α HS.

It is important that the user collect specimens in accordance with the specific assays to be performed. Please see the manufacturer’s instructions (EMD Millipore).  Plates can be designed via the EMD Millipore web site found here. Please be sure to select magnetic bead based assays.

All samples should be submitted on dry ice in ready-to-assay form. Any pre-extraction or sample preparation procedures including dilutions are the sole responsibility of the user.

Samples should be submitted in a minimum-binding 96-well plate (we recommend Fisher part number 12-565-505) following a standard assay design layout and accompanied by a template key outlining the sample order. Please email the sample key as well.

Please provide sufficient volume for all samples to be assayed in duplicate, plus an additional 15-30 µL. For a typical assay this would be (2 x 25 µL) + 30 µL = 80 µL. Please check individual plate requirements in case sample volume varies from this example.

Please carefully assess all assay plate requirements before collecting and processing your samples for delivery.

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

The One-Carbon Metabolism SNP Chip Microarray is the Infinium Expanded Multi-Ethnic Genotyping Array (MEGAEX®) array (illumina) containing over 2 million human SNPs.

Please Note: To download an Excel file containing all 2MM SNPs, go here.

We also run a customized version of this array that contains an additional 800 SNPs focused on the 1-carbon pathway.

Send DNA only, however if you only have blood or other body fluids and you need to have your DNA extracted, see our Genomic DNA Extraction recharge rate.

Procedure for Preparing DNA for the illumine MEGAEX® Array

Extract DNA with any standard method such as phenol-chloroform or a column-based method. We recommend a column-based method such as Qiagen’s QIAamp DNA Blood Mini Kit (Qiagen). The column methods are efficient, automatable, and have less chance of yielding samples with contaminants that could inhibit downstream applications. Per UNC Genotyping Core, DNA must be dissolved in 10 mM Tris-HCL pH 8.0, 1 mM EDTA (TE) (equivalent to Qiagen’s buffer AE).

Make sure that the DNA is of high quality (A260/280 ratio of ~1.8; a gel should be run to check integrity) and free from contaminating reagents (such as ethanol or phenol). To ensure the highest possible quality DNA, a column based method is recommended. Quantitate DNA using the Quant-iT™ PicoGreen ® dsDNA Kit (Invitrogen # P11496).

DNA Preparation Requirements for MEGA-EX SNP Genotyping:

  • Send samples in a 96 well plate (8 by 12).
  • Concentration*:  50 µl of DNA at a concentration of 50-100 ng/µl (2.5-5 µg total DNA)
  • Dilute purified genomic DNA in 10 mM Tris-HCL pH 8.0, 1 mM EDTA (TE).
  • Prepare a sample sheet with this information:  PLATE Name, POSITION, Sample Id, A260, A280, 260/280 ratio, DNA conc (ng/µL), DNA volume in plate (µL), and Total DNA amount (ng)
  • A sample sheet template will be provided.
  • One sample sheet for all the plates. Comma separated CSV, Excel or tab delimited txt format is acceptable.
  • Samples must be coded to ensure anonymity.
  • A copy of the IRB under which the samples were collected is required and methods must include processing and genotyping of samples.
  • Include an MGC Material Transfer Agreement signed by the PI named in your IRB approval.  MGC Material Transfer Agreement can be downloaded at:

Samples not following the above requirements will not be processed.

* Illumina recommends the Molecular Probes PicoGreen® assay for quantitation of dsDNA samples. We will use NanoDrop ND-1000 Spectrophotometer to double-check the DNA concentration.

Materials Transfer Agreement-Mammalian Genotyping Core

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

Service Description

Metabolic Molecular Phenotyping Core

The assay for the Organic Choline Metabolites phosphatidylcholine and sphingomyelin includes analysis of plasma, serum, urine, breast milk, liquid, food, and tissues.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

Service Description

Metabolic Molecular Phenotyping Core

Purified genomic DNA from blood or tissues is analyzed by PCR for Single Nucleotide Polymorphisms (SNPs) for One Carbon Metabolism, (e.g.: PEMT rs12325817, CHDH rs12676 and CHDH rs9001). While we specialize in these SNPs, others can be analyzed by special request. There may be an extra charge for reagent purchase for special request SNPs.

In most cases, TaqMan® SNP Genotyping Assays will be used to detect SNPs and real time PCR carried out on an Eppendorf Mastercycler ep gradient Realplex4 (Eppendorf).

If you need DNA extracted from your sample, see ‘Genomic DNA Extraction’ service for details. If you are extracting your own DNA, we recommend a column-based method such as Qiagen’s QIAamp DNA Blood Mini Kit (Qiagen). Column methods are efficient, automatable, and have less chance of yielding samples with contaminants that could inhibit downstream applications. DNA should be dissolved in 10mM Tris-HCL pH 8.0, 1mM EDTA (TE) (equivalent to Qiagen’s buffer AE). DNA must be of high quality (A260/280 ratio of ~1.8) and free from contaminating reagents such as ethanol or phenol. To ensure the highest possible quality DNA, a column based method is strongly suggested. Quantitate DNA using the Quant-iT™ PicoGreen ® dsDNA Kit (Invitrogen # P11496).

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

This assay is designed to measure Phospholipid Fatty Acids in plasma, breast milk, food or tissues. The following phospholipid fatty acids can be measured by GC-FID:

FA Name n-x Common Name
14:0 Myristic acid
16:0 Palmitic acid
9Trans 16:1 n-7 Palmitelaidic acid
Cis 16:1 n-7 Palmitoleic acid
18:0 Stearic acid
9Trans 18:1* n-9 Elaidic acid
11Trans 18:1 n-7 Vaccenic acid
18:1 n-9 Oleic acid
9T, 12T 18:2 n-6 trans-9,trans-12-Octadecadienoic acid
9C, 12T 18:2 n-6 cis-9,trans-12-Octadecadienoic acid
9T, 12C 18:2 n-6 trans-9,cis-12-Octadecadienoic acid
9C, 12C 18:2 n-6 cis-9,cis-12-Octadecadienoic acid
20:0 Arachidic acid
18:3 n-3 alpha-Linolenic acid (ALA)
20:1 n-9 11-Eicosenoic acid
20:2 n-6 11,14-Eicosadienoic acid
22:0 Behenic acid
20:3 n-6 8,11,14-Eicosatrienoic acid (DHGLA)
20:4 n-6 Arachidonic acid
22:1 n-9 Erucic acid
20:5 n-3 Eicosapentaenoic acid (EPA)
24:1 n-9 Nervonic acid
22:5 n-3 Docosapentaenoic acid (DPA)
22:6 n-3 Docosahexaenoic acid (DHA)

All samples are extracted once and injected twice (see the SOP) for details).

Please note that some phospholipid fatty acids in serum or plasma are represented at very low levels that are at, or near, the limit of detection by the GC-FID method. As such, the %cv for these is significantly higher than that for the more highly represented fatty acids.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

Service Description

Metabolic Molecular Phenotyping Core

Training on the Seahorse XF24 Analyzer (Seahorse Biosciences) is a 1-2 day, hands-on course where you will learn how to setup a protocol and run a plate of cells to measure basal respiration, also known as the oxygen consumption rate (OCR), and the extracellular acidification rate (ECAR). OCR and ECAR are measured simultaneously in each well of the plate by special solid state sensor probes. The trainee will grow their cells of interest and plate them at various densities to determine the optimal seeding density (typically between 20,000 and 60,000 cells/well). There should be a linear increase in basal OCR with cell density. An optimal OCR is from 100 to 400 pmol/min. ECAR will plateau at higher cell densities, so the optimal ECAR is at a seeding density that will allow the user to observe both increases or decreases in ECAR (and OCR) in response to a cell treatment or genetic manipulation.

The trainee will learn how to process the data and how to design future experiments in their cell of choice. Potential trainees should visit the Seahorse Bioscience website to become familiar with the system. Seahorse provides the XF Cell Mito Stress Test for analysis of mitochondrial function, and the XF Glycolysis Stress Test for analysis of glycolytic function, however these are not part of the training.

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

Service Description

Metabolic Molecular Phenotyping Core

Assay for trimethylamine (TMA), trimethylamineoxide (TMAO), choline or betaine in plasma, urine, or tissue.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

Service Description

Metabolic Molecular Phenotyping Core

Cell sorting and flow cytometry can be performed by users trained specifically on the Aria III (BD Biosciences) Fluorescence Activated Cell Sorter (FACS). Sorts can be optimized for purity, yield or single cell modes. The Aria III is equipped with blue (488 nm), green (561 nm), red (633 nm) and violet (405 nm) lasers with 11 detectors that can measure up to 10 colors simultaneously. The hourly rental rate includes sheath fluid, calibration (CS&T) beads, Accudrop beads and other standard solvents used to run and maintain the FACS.

Standard Operating Procedures

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

 

 

Service Description

Metabolic Molecular Phenotyping Core

The Zeiss Axio Imager A1 Microscope and Camera is available for hourly rental. This microscope has brightfield, darkfield, differential phase interference contrast (DIC), polarization contrast (PO) and fluorescence capabilities. Available filter sets for DAPI, FITC and rhodamine detection. AxioVision 4.8.2 software available on attached pc. The scope is located in room 2341 of the Nutrition Research Institute.

Other Important Information

Information including sample collection instructions, shipping directions, and the standard operating procedures for this assay, can be found on the core’s services web site.

Have Questions? Contact Us.

Steve Orena, Lab Manager-Kannapolis Facilities
Email • Phone: (704) 250-5041

Washington University, St. Louis

Service Description

Biomolecular Analysis Core

Access to state-of-the-art mass spectrometry analyses needed to quantify and/or identify the structure of nutrition/obesity – related biomolecules through a centralized, standardized, cost-effective, and efficient “one-stop-shop.” This includes:

  • Structural identification or quantitation of complex lipids
  • Fatty acids identification or quantitation
  • Sterols, including cholesterol and its oxidation products
  • Low molecular weight metabolic substrates
  • Polyols
  • Amino acids and their oxidation products
  • [13C] or [2H] or [15N] or [18O] enrichment in mixtures of biological substrates and their end-products after heavy isotope-labeled precursor administration
  • Peptide sequencing, post-translational modifications, and assistance with protein identification (MASCOT database searches)
  • Select plant products
  • Assistance preparing biological samples for mass spectrometry analyses (extraction, purification, derivatization) and interpreting mass spectra
  • Assistance in identifying unknown biomolecules
  • Training in operating and maintaining mass spectrometry systems
  • Other nutrition-related biomolecules

Have Questions? Contact Us.

Dan Ory, MD, Core Director
Email • Phone: (314) 362-8737