Enzyme Assay Services
Quantitative 15-Lipoxygenase Activity Assay for Drug Discovery
Validate enzyme function and screen modulators with a reliable, quantitative measurement of 15-lipoxygenase (15-LOX) activity. Our service provides recombinant expression, purification, and activity-based characterization of mammalian arachidonic acid 15-lipoxygenases—key targets in inflammatory and metabolic diseases. Designed for drug discovery groups in biotech and pharma investigating lipoxygenase inhibitors for conditions such as asthma and atherosclerosis.
What Is a 15-Lipoxygenase Activity Assay?
15-Lipoxygenase (15-LOX) catalyzes the dioxygenation of polyunsaturated fatty acids such as arachidonic acid and linoleic acid, producing lipid hydroperoxides that serve as signaling molecules in inflammatory and metabolic pathways. Quantitative measurement of 15-LOX activity is essential for validating enzyme function, characterizing recombinant preparations, and screening potential inhibitors in drug discovery programs.
Our 15-lipoxygenase activity assay service provides a complete workflow—from recombinant protein expression and purification to functional characterization and inhibitor screening. The assay detects lipid hydroperoxides formed by 15-LOX activity, using either fluorometric or colorimetric readouts that are adaptable to purified enzymes and cell lysates. This enables reliable, quantitative assessment of enzyme activity in formats suited to medium-throughput screening.
The service is particularly relevant for drug discovery groups targeting lipoxygenase pathways in conditions such as asthma, atherosclerosis, and other inflammatory diseases, where 15-LOX plays a central role in lipid mediator biosynthesis.
Fluorometric Readout
Fluorescent intensity assay for the detection of lipid hydroperoxides formed by human 15-lipoxygenase in enzyme or cellular assays.
- Sensitive detection of hydroperoxide products
- Applicable to enzyme and cell-based formats
- Suitable for kinetic measurements
Colorimetric Readout
Colorimetric assay adaptable for purified lipoxygenases and cell lysates containing active enzyme, performed in a 96-well format.
- 96-well format for medium-throughput
- Works with purified LOXs and lysates
- Simple spectrophotometric measurement
Inhibitor Screening
Assay formats suited for measuring inhibition concentration 50% (IC50) and enzyme kinetic parameters for modulator screening.
- IC50 determination
- Enzyme kinetic analysis
- Medium-throughput screening capability
Why Measure 15-LOX Activity?
15-Lipoxygenase is a key enzyme in the biosynthesis of pro-inflammatory and pro-resolving lipid mediators. Dysregulation of 15-LOX activity has been implicated in inflammatory diseases, atherosclerosis, and metabolic disorders, making it an attractive target for therapeutic intervention. Reliable activity measurement is critical for validating enzyme function, assessing the impact of genetic or pharmacological modulation, and progressing inhibitor programs.
The assay detects the increase in absorbance at 234 nm resulting from the conjugation of double bonds in fatty acid hydroperoxides formed by 15-LOX-catalyzed dioxygenation. This well-established spectrophotometric principle provides a quantitative basis for comparing enzyme activity across conditions, inhibitors, and preparations.
| Application | Readout | Format | Use Case |
|---|---|---|---|
| Enzyme validation | Fluorometric / colorimetric | Purified enzyme | Confirm activity of recombinant 15-LOX |
| Cell lysate analysis | Colorimetric | 96-well plate | Measure endogenous or overexpressed 15-LOX |
| Inhibitor screening | Fluorometric | 96-well plate | Determine IC50 of candidate modulators |
| Kinetic characterization | Fluorometric / colorimetric | Purified enzyme | Assess substrate affinity and catalytic parameters |
How the Service Works
The 15-lipoxygenase activity assay service follows a structured workflow designed to deliver reliable, quantitative data for your enzyme characterization and screening needs. Each step is performed with attention to reproducibility and quality control.
Project Consultation
We discuss your specific research goals, whether enzyme validation, inhibitor screening, or kinetic characterization. We confirm the appropriate 15-LOX isoform, assay format, and sample requirements for your study.
Recombinant Expression and Purification
Mammalian arachidonic acid 15-lipoxygenases are expressed in a suitable recombinant system and purified to yield active enzyme preparations suitable for activity assays.
Activity Assay Setup
The assay is configured using either fluorometric detection of lipid hydroperoxides or colorimetric measurement of conjugated dienes at 234 nm, depending on your requirements and sample type.
Data Analysis and Reporting
Raw data are analyzed to calculate enzyme activity, kinetic parameters, or inhibition values. A detailed report documents the methods, results, and quality control measures applied throughout the study.
Customization Options
Every research program has unique requirements. Our 15-lipoxygenase activity assay service can be tailored to match your specific experimental design, sample types, and data needs.
Whether you need to screen a focused set of compounds, characterize a novel enzyme variant, or compare activity across cell lines, the service can be adapted to deliver the information most relevant to your program.
Assay Format Selection
Choose between fluorometric and colorimetric readouts based on sensitivity requirements and available instrumentation.
- Fluorometric for higher sensitivity
- Colorimetric for simple spectrophotometric detection
- 96-well format for medium-throughput
Sample Type Compatibility
The assay is adaptable to purified enzymes and cell lysates containing active 15-LOX, accommodating diverse experimental systems.
- Purified recombinant enzyme
- Cell lysates with endogenous or overexpressed enzyme
- Optimized protocols for each sample type
Screening and Kinetics
Configure the assay for inhibitor screening with IC50 determination or for detailed enzyme kinetic characterization.
- IC50 measurement for candidate inhibitors
- Kinetic parameter determination
- Medium-throughput screening capability
Deliverables and Documentation
Each project concludes with a comprehensive report that enables you to interpret results with confidence and use the data in downstream decision-making.
The documentation is designed to support regulatory and publication needs, providing transparency into the methods and quality controls applied.
| Deliverable | Description | Format | Use |
|---|---|---|---|
| Activity data | Quantitative measurement of 15-LOX activity | Excel / PDF | Enzyme validation and comparison |
| Kinetic parameters | Calculated kinetic constants where applicable | Excel / PDF | Enzyme characterization |
| Inhibition data | IC50 values for screened compounds | Excel / PDF | Hit identification and lead optimization |
| Method report | Detailed protocol and quality control documentation | Publication and regulatory support |
Quality and Reproducibility
Reliable enzyme activity data depend on careful assay design and rigorous quality control. Our service applies established methods for lipoxygenase activity measurement, with attention to reproducibility across runs and batches.
Quality control measures include verification of enzyme activity with positive controls, appropriate blanks, and consistent assay conditions to ensure that results are comparable across experiments.
Positive Controls
Assay performance is verified using enzyme preparations with known activity to confirm the assay system is functioning correctly.
- Known-active enzyme controls
- Confirmation of assay sensitivity
- Batch-to-batch consistency
Reproducible Conditions
Standardized assay conditions including substrate concentration, buffer composition, and temperature are maintained across experiments.
- Defined substrate and buffer conditions
- Consistent incubation parameters
- Comparable results across runs
Data Verification
Raw data are reviewed to confirm the validity of measurements before analysis and reporting.
- Review of raw absorbance/fluorescence values
- Verification of linearity where applicable
- Documentation of any anomalies
Applications in Drug Discovery
15-Lipoxygenase is a validated target in inflammatory and metabolic diseases, and reliable activity assays are essential for advancing inhibitor programs. The service supports multiple stages of the drug discovery pipeline.
From target validation to hit identification and lead optimization, quantitative 15-LOX activity data provide the foundation for informed decision-making.
| Stage | Application | Data Provided | Decision Impact |
|---|---|---|---|
| Target validation | Confirm 15-LOX activity in disease-relevant models | Activity levels in relevant systems | Validate target engagement |
| Hit identification | Screen compound libraries for 15-LOX inhibition | IC50 values for hits | Prioritize compounds for further study |
| Lead optimization | Compare potency of analogs and series | Relative inhibition data | Select lead candidates |
| Mechanism of action | Assess inhibition mode and selectivity | Kinetic and inhibition profiles | Understand compound behavior |
Why Choose This Service
The 15-lipoxygenase activity assay service combines established detection methods with a flexible, project-focused approach. Whether you need to validate enzyme function, screen inhibitors, or characterize kinetics, the service is designed to deliver reliable data efficiently.
By providing a complete workflow from recombinant expression to activity measurement, the service removes the need for specialized in-house assay setup and allows your team to focus on interpretation and decision-making.
Complete Workflow
From recombinant expression and purification to activity assay and data reporting, the service covers the full process.
- No need for in-house assay setup
- Integrated expression and characterization
- Single point of contact
Established Methods
The assay uses well-documented approaches for lipoxygenase activity measurement, including fluorometric and colorimetric detection.
- Spectrophotometric detection at 234 nm
- Fluorometric detection of hydroperoxides
- Published and validated methodologies
Flexible Formats
Assay configurations can be adapted to purified enzymes or cell lysates, and to screening or kinetic studies.
- Purified enzyme or lysate compatibility
- 96-well format for medium-throughput
- IC50 and kinetic determinations
Service Comparison
The 15-lipoxygenase activity assay service offers a comprehensive solution compared to common alternatives. The table below highlights key differentiators.
FAQ
What sample types can be used for the 15-LOX activity assay?
The assay is adaptable to both purified lipoxygenases and cell lysates containing active enzyme. This flexibility allows measurement of recombinant enzyme preparations, endogenous enzyme in cell lines, or overexpressed 15-LOX in transfected cells. The appropriate format is confirmed during project consultation to ensure compatibility with your sample type.
How is 15-LOX activity measured in this assay?
The assay detects lipid hydroperoxides formed by 15-lipoxygenase activity. Two detection options are available: a fluorometric intensity assay for sensitive detection of hydroperoxides, and a colorimetric assay based on the increase in absorbance at 234 nm resulting from conjugated diene formation. The choice of readout depends on your sensitivity requirements and available instrumentation.
Can the service be used for inhibitor screening?
Yes, the assay format is suited for medium-throughput screening and can be configured to measure inhibition concentration 50% (IC50) for candidate modulators. This makes the service applicable to hit identification and lead optimization programs targeting 15-lipoxygenase in inflammatory and metabolic diseases.
What data will I receive at the end of the project?
You will receive a comprehensive report including quantitative activity data, calculated kinetic parameters where applicable, and inhibition data for screened compounds. The report documents the methods used and quality control measures applied, providing transparency for publication and regulatory purposes.
References
- Crafting Molecular Tools for 15‐Lipoxygenase‐1 in a .. by A Louka · 2025 · Cited by 7 — Workflows for competition and irreversibility assays to assess in cellulo target engagement of 15-LOX-1 by BAM10, using confocal microscopy. View article
- Dzoyem JP, Kuete V, McGaw LJ. The 15-lipoxygenase inhibitory, antioxidant, antimycobacterial activity and cytotoxicity of fourteen ethnomedicinally used African spices and culinary herbs. J Ethnopharmacol 2014 Oct 28. View on PubMed
- by W Lu · 2013 · Cited by 67 — The assay is adaptive for purified LOXs and cell lysates containing active LOXs. Development of a new colorimetric assay for lipoxygenase .. We use the new colorimetric assay in a 96-well format to ... View on PubMed
- Determination of Lipoxygenase Activity in Plant Extracts Using a .. Typically, the methodologies used for the measurement of lipoxygenase activity are the spectrophotometric assay based on the measurement of the absorbance at ... View article
- Briefly, the activity of 15-lipoxygenase is observed as it catalysed the reaction between oxygen and linoleic acid. 15-Lipoxygenase inhibition of Commelina benghalensis ... - PMC. The increase in absorbance at 234 nm was due ... View article
Advance Your 15-LOX Program
Contact us to discuss how the 15-lipoxygenase activity assay service can support your enzyme validation, inhibitor screening, or kinetic characterization needs. Our team will work with you to design a project that delivers the data you need.