Cancer Metabolism (La Jolla)

Services

The primary technology and tools in the La Jolla core are:

• GC/MS-based analysis of metabolites such as amino acids, keto acids and lipids. Measuring both the abundances of these metabolites and the rate of stable isotope labeling enables determination of the relative activity of a number of metabolic pathways involved in tumor progression, including glycolysis, the pentose phosphate pathway, the TCA cycle, de novo fatty acid biosynthesis and amino acid biosynthesis. In addition, the metabolism or exchange of other isotope labeled metabolites can be tracked as warranted.
• Measurement of cellular respiration and glycolysis using a Seahorse XF24. This instrument determines the rate of glycolysis and respiration of adherent cells in a 24-well format. Glycolysis is observed via the acidification of the tissue culture media. Respiration is observed by measuring total oxygen consumption. With proper experimental design, the rates of basal respiration, ATP production, proton leak, maximal respiration, spare respiratory capacity and non-mitochondrial oxygen consumption may be determined.
• Rapid and accurate measurement of major metabolites (glucose, glutamine, lactate, glutamate, ammonium and potassium) using the YSI 7100 analyzer. This instrument is able to rapidly (~1 min per metabolite) determine the concentration of these important metabolism substrates from a small volume of sample (~200 µL).

In addition, the core has access to LC/MS and NMR instrumentation and experts in Sanford-Burnham, La Jolla, LC/MS/MS at the Lake Nona site, with broad, unbiased metabolite profiling available locally on a fee-per-samples basis in the Scripps Center for Metabolomics and Mass Spectrometry (La Jolla).

Equipment and Resources

• The YSI 7100 metabolite analyzer is able to measure glucose, glutamine, lactate, glutamate, ammonia and potassium in media or other liquid samples. Amenable to most concentrations found in laboratory samples. Requires 10 - 25 µL of sample per injection/assay. Due to the nature of the equipment it is most cost effect to run many samples at the same time or within the same week.
• The Seahorse XF24 can simultaneously measure the two major energy producing pathways of live cells (mitochondrial respiration and glycolysis) in a microplate in real-time. This fast and sensitive measurement of cellular bioenergetics is label free, enabling time-resolved analysis and the reuse of the cells.
• The Shimadzu QP2010 gas chromatograph-mass spectrometer is used for broader metabolic flux analysis including determination of stable isotope labeling rates of various intra- and extracellular metabolites, or determining the abundance of amino, keto and fatty acids. It has an expanded mass range (1.5 to 1090 Daltons), uniform temperature control, and an expanded temperature range of 100 to 300°C.
• A Shimadzu Prominence HPLC is available for the measurement of metabolites not generally accessible by GC/MS. In particular, assays have been developed to detect ATP, ADP, AMP and NAD species. The HPLC is also ideal for bulk amino acid analysis via automated HCl hydrolysis. Detectors include a dual UV/Vis , fluorescence and refractive index. Both the column and autosampler are temperature controlled.

Additional Sanford-Burnham Analytical Resources Available for Metabolomics

• Lake Nona Metabolomics Core - Utilizes the Duke Steadman modules for analysis of Acylcarnitines, Organic acids, and Amino acids. The core is working to expand its repertoire to include focused analysis, and very broad Orbitrap-based profiling.
• Lake Nona (in vivo) Metabolic Phenotyping Core – CLAMS metabolic cage system (food, water, excretion, O2/CO2), exercise metabolism, NMR body composition analysis, and extensive glucose/insulin studies.
• NMR-based analysis - Can be accessed in La Jolla through collaboration with J. Smith Lab and the NMR Facility.
• Lipidomics - Shotgun lipidomics, in collaboration with X. Han lab at Lake Nona.
• Seahorses (both XF24 and 96) – Common equipment at Lake Nona.
• Other LC-MS Instruments in La Jolla – API 3000 LC/MS/MS, and several other LC/MS systems in the Proteomics core facility.
• Informatics core in Lake Nona with particular expertise in analysis of metabolic data, and integration of these data with other data types such as genomics.

External Resources

• Scripps LC-MS/MS facility - (Center for Metabolomics and Mass Spectrometry) for broad profiling of metabolites.

Additional Sanford-Burnham Analytical Resources Available for Metabolomics

Analytical

• Lake Nona Metabolomics Core - Utilizes the Duke Steadman modules for analysis of Acylcarnitines, Organic acids, and Amino acids. The core is working to expand its repertoire to include focused analysis, and very broad Orbitrap-based profiling.
• Scripps LC-MS/MS facility for broad profiling (fairly inexpensive and produces much data, but large datasets challenging to interpret)
• Lake Nona (in vivo) Metabolic Phenotyping Core – CLAMS metabolic cage system (food, water, excretion, O2/CO2), exercise metabolism, NMR body composition analysis, and extensive glucose/insulin studies.
• NMR-based analysis - Can be accessed in La Jolla through collaboration with J. Smith Lab.
• Lipidomics - Shotgun lipidomics technology in Xianlin Han’s lab at Lake Nona, currently available through collaboration with Han lab
• Seahorses (both XF24 and 96) – Common equipment at Lake Nona, accessible through collaboration
• Other LC-MS Instruments in La Jolla – In Med Chem, there is an available API 3000 LC/MS/MS, and several other LC/MS systems there and in Proteomics

Computational

• Informatics core staff – In Lake Nona, Feng Qi, PhD, was hired for expertise in modeling metabolomics data, as well as integrating these data with other data types
• Duke Modules – The computational modules from Steadman essentially deconvolutes LC-MS peaks into identified metabolites

Contact

Adam D. Richardson, Ph.D.
Director, Cancer Metabolism Resources

• Email: adrichardson@sanfordburnham.org
• Office: 858.795.5136 (or x5136)

David Scott, Ph.D.
Staff Scientist

• Email: dscott@sanfordburnham.org
• Office: 858.795.3941 (or x3941)

Jorge Moscat, Ph.D.
Faculty Director

• Email: jmoscat@sanfordburnham.org

Price List