Metabolomics Facility

Techniques

Most biological processes, including energy production, growth, waste disposal and gene expression, require proper control of metabolism. Normal tissue function and development cannot occur without precise control of metabolism, and as a result, many diseases involve perturbed metabolism at the cellular level. Such diseases include inborn errors of metabolism caused by inherited mutations in metabolic enzymes and more common, multifactorial diseases like cancer, diabetes and cardiovascular disease.

Our facility uses an array of techniques to assess metabolism.

These techniques include:

Isotope enrichment analysis
These techniques involve introduction of a nutrient labeled with a stable isotope (e.g. ¹³C, ¹⁵N, ²H) into a biological system, followed by measurement of isotope enrichment in metabolites downstream of the labeled nutrient tracer, to assess flow through metabolic pathways. The choice of instrument depends on the metabolites to be analyzed. Core personnel are available to assist in study design.

• Yang et al., 2014, Mol Cell, 56, 414-424
• Eskiocak et al., 2016, Nat Commun, 7, 12336
• Faubert et al., 2017, Cell, 171, 358-371
• Tasdogan et al., 2020, Nature, 577, 115-120

Targeted metabolomics analysis
These assays use tailored mass spectrometry approaches to assess the levels of specific metabolites pre-chosen by the user or the Core. These may include single metabolites for absolute quantitation (“Metabolite Quant”), several metabolites chosen to assess a single pathway (“Pathway Analysis”) or “Screening Analysis” using assays developed by the Core to detect hundreds of metabolites simultaneously.

• Gu et al., 2019, Cancer Discov, 9, 1228-1247
• Tasdogan et al., 2020, Nature, 577, 115-120

Screening Analyses are the most commonly-requested assays. These assays include a triple-quadrupole method designed to detect ~150 metabolites with optimized sensitivity, and a broader assay on high-resolution mass spectrometry that routinely detects several hundred metabolites. Core personnel are available to assist in choosing the assay that best meets the experimental question.

• Mullen et al., 2014, Cell Reports, 7, 1679-1690
• Piskounova et al., 2015, Nature, 527, 186-191
• Tasdogan et al., 2020, Nature, 577, 115-120

Untargeted metabolomics analysis
This is a shotgun-like approach that acquires all possible metabolite data from experimental samples with no prioritization of which pathways might be informative. The method reports the maximum number of metabolic features (i.e. “peaks”) and identifies as many as possible by comparing the precise molecular mass to commercial databases. For a large number of peaks, a precise mass but not a definitive metabolite identification will be available.

Seahorse assay
Measures oxygen consumption and extracellular acidification in a 96-well format from a variety of sample types, including cultured cells and isolated mitochondria.

• Sapir et al., 2014, Proc Natl Acad Sci USA, 111, e3880-9
• Zhang et al., 2017, eLife, 6:e17834
• Cappel etal., 2019, Cell Metab, 29, 1291-130
• Lesner et al., 2020, Metab Eng, 60, 157-167
• Chen et al., 2020, eLife, 29;9:e58108