Metabolic Pathway Assays and Functional Profiling

CD Biosynsis offers advanced Metabolic Pathway Assays essential for understanding cellular bioenergetics, nutrient utilization, and drug mechanism of action in areas such as cancer metabolism, diabetes, and aging. Metabolic reprogramming is a hallmark of many diseases, making key metabolic enzymes and transporters attractive therapeutic targets. Our platform provides quantitative analysis of pathway flux, enzyme activity, and metabolite concentrations. We utilize state-of-the-art technologies including high-resolution respirometry, enzyme-linked colorimetric/fluorometric assays, and targeted mass spectrometry (LC-MS/MS) for stable isotope tracing. Researchers can select and combine individual pathway modules to build a custom profiling panel, enabling precise characterization of compound effects on critical cellular processes like glycolysis, oxidative phosphorylation, and fatty acid oxidation.

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Quantitative Flux Analysis and Enzyme Characterization

Metabolic pathways are interconnected, and perturbing one step can have cascading effects. Reliable drug screening requires precise tools to measure both global cellular output (e.g., oxygen consumption rate) and specific enzymatic bottlenecks (e.g., pyruvate kinase activity). Our service leverages specialized platforms, such as the Seahorse XF Analyzer for bioenergetics and targeted LC-MS/MS for absolute metabolite quantification. By offering selectable modules, you gain the flexibility to focus on specific pathways, such as assessing compound effects on the Warburg effect (aerobic glycolysis) or characterizing the potency of inhibitors against key enzymes like IDH or mitochondrial complex components. This approach provides clear, quantitative evidence of drug impact on cellular energy metabolism.

Selectable Metabolic Pathways and Enzyme Targets

Core Bioenergetics Pathways Metabolic Enzyme Targets and Inhibitors Advanced Functional Analysis Modules

Core Cellular Bioenergetics Pathways

Choose Global Pathways for Functional Profiling

Check the box next to the Metabolic Pathway you wish to profile for flux or activity changes:

Glycolysis Stress Test (Extracellular Acidification Rate)

Mitochondrial Respiration (Oxygen Consumption Rate)

Fatty Acid Oxidation (FAO) Flux

Glucose Uptake Assay

Pentose Phosphate Pathway (PPP) Flux

Glutamine Metabolism (Glutaminolysis)

TCA Cycle Flux (Citric Acid Cycle)

ATP Production Rate Assay

Metabolic Enzyme Targets and Inhibitors

Choose Specific Enzymes for Activity and IC50 Profiling

Select the specific Metabolic Enzymes you require for activity and inhibitor screening:

IDH1/IDH2 (Isocitrate Dehydrogenase)

LDH (Lactate Dehydrogenase)

HK2 (Hexokinase 2)

Pyruvate Kinase (PKM2)

FASN (Fatty Acid Synthase)

Complex I (Mitochondrial Respiration)

FBPase (Fructose-1,6-bisphosphatase)

All other available targets (Inquire)

Advanced Functional Analysis Modules

Select Mechanistic and Quantitative Services

Stable Isotope Tracing (Flux Analysis)

LC-MS/MS based tracing using 13C-labeled glucose or glutamine to map specific pathway contribution.

Targeted Metabolite Quantification

Absolute quantification of 50+ central metabolites (e.g., NAD+/NADH, Lactate, key amino acids) via LC-MS/MS.

Mitochondrial Function and Toxicity

High-resolution respirometry and membrane potential assays for assessing compound toxicity and coupling efficiency.

Metabolic Pathway Assay Service Workflow

A structured process for comprehensive metabolic profiling.

Panel Selection and Sample Preparation

Bioenergetics and Enzyme Activity Screening

Metabolite Quantification and Flux Analysis

Data Interpretation and Reporting

Panel Customization: Finalize the list of pathways and specific enzyme targets (IC50) based on client selection.

Cell Culture Prep: Optimize cell density and media conditions for consistent metabolic activity across all assays.

Bioenergetics Assay: Run Seahorse XF analysis for OCR and ECAR measurements (Basal, Stress Test, ATP Production).

Enzyme Activity: Screen inhibitors against selected purified metabolic enzymes (e.g., IDH2) to determine IC50 values.

Metabolite Extraction: Quench cell metabolism rapidly and extract polar metabolites for MS analysis.

Isotope Tracing: Incubate cells with labeled substrate (e.g., 13C-Glucose) and measure fractional enrichment of downstream metabolites via LC-MS/MS.

  • Flux Calculation: Determine the relative contribution of different pathways (e.g., Glycolysis vs. TCA) to biomass or energy production.
  • Selectivity Index: Compare IC50 of target enzyme against bioenergetic assay IC50 for MoA correlation.
  • Reporting: Deliver comprehensive data including OCR/ECAR plots, IC50 values, and calculated flux maps.

Precision and Insight in Cellular Metabolism

Targeted Mass Spectrometry

           

High-confidence quantification of central metabolites and precise analysis of stable isotope tracing flux.

Integrated Bioenergetics

           

Full utilization of Seahorse XF technology for real-time measurement of cellular respiration and glycolysis.

Customizable Pathway Panels

           

Flexibility to combine pathway flux analysis with specific enzyme inhibition profiling for MoA clarity.

Enzyme Inhibition Screening

           

Quantitative IC50 and kinetic profiling of inhibitors against purified enzymes of major metabolic pathways.

Client Testimonials on Metabolic Assays

"The stable isotope tracing study was instrumental. It definitively showed that our compound was rerouting glutamine away from the TCA cycle, clarifying its anti-cancer mechanism."

Dr. Jian Li, Cancer Metabolism Research

"The Seahorse XF analysis, combined with the enzyme IC50 profiling, provided a clear, integrated picture of our lead's dual impact on glycolysis and mitochondrial respiration."

Ms. Anya Sharma, Preclinical Development Lead

"Their targeted metabolomics service provided absolute quantification of key NAD+/NADH ratios, essential data that correlated perfectly with our target enzyme's functional readout."

Mr. Julian Chen, Therapeutic Discovery Lead

FAQs about Metabolic Pathway Assays

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What is the difference between OCR and ECAR?

OCR (Oxygen Consumption Rate) is a measure of mitochondrial respiration. ECAR (Extracellular Acidification Rate) is a measure of glycolytic activity, primarily due to the production of lactic acid.

What is the purpose of stable isotope tracing?

Stable isotope tracing (e.g., using 13C-Glucose) measures the flux, or movement, of atoms through metabolic pathways, providing insight into how a pathway contributes to downstream metabolites, which is often superior to simple concentration measurements.

Can you perform assays on primary cells or patient-derived cells?

Yes. Many of our metabolic assays, particularly those using Seahorse XF and targeted MS, are optimized for use with challenging samples, including primary cells and patient-derived xenografts (PDX) in culture.

What is the Warburg Effect?

The Warburg Effect describes the observation that many cancer cells preferentially produce energy via a high rate of glycolysis followed by lactic acid fermentation, even in the presence of sufficient oxygen (aerobic glycolysis).

How much does Metabolic Engineering services cost?

The cost of Metabolic Engineering services depends on the project scope, complexity of the target compound, the host organism chosen, and the required yield optimization. We provide customized quotes after a detailed discussion of your specific research objectives.

Do your engineered strains meet regulatory standards?

We adhere to high quality control standards in all strain construction and optimization processes. While we do not handle final regulatory approval, our detailed documentation and compliance with best laboratory practices ensure your engineered strains are prepared for necessary regulatory filings (e.g., GRAS, FDA).

What to look for when selecting the best gene editing service?

We provide various gene editing services such as CRISPR-sgRNA library generation, stable transformation cell line generation, gene knockout cell line generation, and gene point mutation cell line generation. Users are free to select the type of service that suits their research.

Does gene editing allow customisability?

Yes, we offer very customised gene editing solutions such as AAV vector capsid directed evolution, mRNA vector gene delivery, library creation, promoter evolution and screening, etc.

What is the process for keeping data private and confidential?

We adhere to the data privacy policy completely, and all customer data and experimental data are kept confidential.

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