Services

Professional and Cost-Saving Solutions

  1. Home
  2. Screening of Substrates, Inhibitors, and Other Ligands
  3. Inhibitor Screening and Design
  4. High-Throughput Screening of Inhibitors
  5. High-Throughput Inhibitor Assays and Screening

High-Throughput Inhibitor Assays and Screening

inquiry

Creative Enzymes provides comprehensive High-Throughput Inhibitor Assays and Screening (HTS) services to accelerate the discovery of enzyme inhibitors for pharmaceutical, food, and industrial applications. By combining advanced assay technologies, automation platforms, and expert enzymology, we deliver precise, scalable, and reproducible screening of thousands of compounds. Whether applied to early hit identification or large-scale validation campaigns, our services offer clients a reliable foundation for advancing promising inhibitors into further development.

Understanding High-Throughput Inhibitor Assays and Screening

High-throughput screening is the cornerstone of modern inhibitor discovery. Traditional, small-scale enzymatic assays are not sufficient for handling the vast compound libraries needed in today's drug and biocatalyst development pipelines. HTS offers the ability to evaluate thousands of compounds in parallel, providing both efficiency and statistical robustness.

High-throughput assays for enzyme inhibitor screening

However, the complexity of assay design, automation, and data analysis often presents challenges. Inhibitor screening requires assays that are not only sensitive and reproducible but also compatible with miniaturized, automated workflows. Moreover, specialized expertise is essential for adapting assay conditions to different enzyme families, ensuring that results are biologically relevant.

Creative Enzymes addresses these challenges by integrating deep enzymology expertise with advanced HTS technologies. We offer tailored assays, state-of-the-art automation, and data analysis pipelines to support a broad spectrum of inhibitor discovery programs.

Our Service Offerings

Our High-Throughput Inhibitor Assays and Screening service is designed to support clients through every aspect of inhibitor testing, from assay development to large-scale screening campaigns.

Services We Offer

Assay Development and Optimization

  • Tailored assay formats, including spectrophotometric, fluorometric, luminescent, and absorbance-based readouts.
  • Adaptation of assays for specific enzyme families (kinases, proteases, glycosidases, lipases, esterases, transferases, etc.).
  • Validation of dynamic range, signal-to-noise ratio, and reproducibility.

High-Throughput Screening (HTS)

  • Screening of thousands of compounds using 96-, 384-, or 1536-well microplate formats.
  • Automated liquid handling systems for consistent sample preparation.
  • Integration of robotics, plate readers, and data capture systems for efficient workflows.

Assay Formats and Strategies

  • Biochemical Assays: Monitoring substrate turnover, product formation, or cofactor consumption.
  • Binding Assays: Using surface plasmon resonance (SPR), fluorescence polarization (FP), or biolayer interferometry (BLI).
  • Mechanism-Based Assays: Designed to distinguish between competitive, non-competitive, or allosteric inhibitors.
  • Stability and Specificity Testing: Inhibitor activity under varied pH, temperature, or solvent conditions; assessment of off-target interactions.

Data Analysis and Deliverables

  • Raw and processed assay data in standardized formats.
  • Quantitative measurements of inhibition (IC50, Ki, dose–response curves).
  • Hit identification with statistical confidence measures (Z′ factor, signal window).
  • Ranked lists of candidate inhibitors with recommendations for follow-up studies.
  • Comprehensive final report including assay design, conditions, and detailed results.

With this integrated approach, Creative Enzymes provides clients with not only high-quality data but also actionable insights for downstream optimization.

Contact Our Team

Our Comprehensive Services for High-Throughput Inhibitor Screening

In addition to the screening itself, we provide a full suite of supporting services to ensure reliable results and seamless project progression:

Inquiry

Our Advantages

Broad Assay Expertise

Capability to design assays across diverse enzyme families and detection technologies.

Advanced Automation

Robotic liquid handling and plate readers ensure high precision and throughput.

Scalable Formats

From small pilot screens to full-scale campaigns with thousands of compounds.

Data Quality Assurance

Rigorous validation steps guarantee sensitivity, reproducibility, and low error rates.

Customizable Solutions

Flexible workflows tailored to client targets, compound libraries, and assay preferences.

End-to-End Integration

Smooth connection with library construction, activity measurement, and SAR studies.

Case Studies and Real-World Applications

Case 1: Discovery of Potent SARS-CoV-2 Mpro Inhibitors

The main protease (Mpro) of SARS-CoV-2 is a critical antiviral target. A new fluorogenic substrate was developed for high-throughput screening, outperforming conventional fluorescent peptides. Using focused libraries, two inhibitor classes—peptidomimetic azanitriles and pyridyl indole esters—were identified, optimized, and biochemically characterized. Azanitrile derivatives simultaneously inhibited Mpro and cathepsin L, relevant for viral entry, while pyridyl esters were evaluated via positional scanning. Two irreversible inhibitors, azanitrile 8 (Ki = 24 nM) and pyridyl ester 17 (Ki = 10 nM), showed exceptional potency. These rationally designed molecules represent promising candidates for further antiviral development, highlighting the efficacy of targeted library approaches over conventional virtual screening.

Development of SARS-CoV-2 main protease inhibitors through high-throughput screening and rational designFigure 1. The main protease (Mpro) of SARS-CoV-2 has been recognized as a significant drug target to treat COVID-19. The design of a new fluorogenic substrate, the recombinant expression of Mpro and the development of an HTS assay were combined with a structure-based rational approach leading to the discovery of tailored azanitriles and indole pyridyl esters as novel, outstandingly potent Mpro inhibitors. (Breidenbach et al., 2021)

Case 2: Liquid Crystal-Assisted Screening of Xanthine Oxidase Inhibitors

Small-molecule inhibitors are essential for therapeutic development, and efficient screening methods are critical. A novel liquid crystal (LC)-based assay was developed, using enzyme catalysis-induced aptamer release to identify xanthine oxidase (XOD) inhibitors. XOD-mediated xanthine oxidation prevents aptamer binding, producing bright LC images, whereas inhibition restores aptamer binding, resulting in dark images. Screening a small library of triazole derivatives identified three potent inhibitors. Molecular docking confirmed active site occupation, and the compounds exhibited excellent biocompatibility in HEK293 and HeLa cells. This high-throughput, label-free LC-based strategy offers high sensitivity and accuracy, providing a promising approach for efficient enzyme inhibitor discovery.

Liquid crystal-based screening assay for xanthine oxidase (XOD) inhibitorsFigure 2. Screening of the potential XOD inhibitors. (A) Chemical structures of selected potential inhibitors. (B) Activity of XOD after inhibition by different concentrations of compound I. (C) Molecular docking of XOD with compound I. The main amino acid residues of XOD in the active site are represented with thin sticks. The violet dashed lines stand for hydrogen bonds, and the hydrogen bond distances are 3.0, 3.1, and 2.7 Å, respectively. (Wu et al., 2021)

FAQs About High-Throughput Inhibitor Assays and Screening

  • Q: How many compounds can you screen in a single campaign?

    A: Our platform supports screening campaigns ranging from a few hundred compounds to over 100,000, depending on the client's needs and library format.

  • Q: What detection methods are available for assays?

    A: We support a wide variety of detection methods, including absorbance, fluorescence, luminescence, fluorescence polarization, and label-free approaches such as SPR and BLI.

  • Q: How do you ensure assay reproducibility and accuracy?

    A: We validate assays prior to screening, applying statistical parameters such as Z′ factors, and incorporate positive/negative controls in every run. Automated systems reduce manual variability, ensuring consistent performance.

  • Q: Can assays be customized for specific enzymes or conditions?

    A: Yes. We specialize in adapting assays to client-defined enzymes, substrates, and conditions (e.g., pH, temperature, cofactors), ensuring biological relevance and industrial applicability.

  • Q: How are false positives minimized during HTS?

    A: We include stringent controls, replicate testing, and orthogonal confirmatory assays to eliminate artifacts. Data analysis pipelines are designed to flag inconsistent or low-confidence results.

  • Q: What deliverables can I expect at the end of a screening project?

    A: Clients receive raw data, processed datasets, dose–response curves, ranked inhibitor lists, and a detailed report summarizing assay design, conditions, and validated hits, along with recommendations for follow-up studies.

References:

  1. Breidenbach J, Lemke C, Pillaiyar T, et al. Targeting the main protease of SARS-CoV-2: from the establishment of high throughput screening to the design of tailored inhibitors. Angew Chem Int Ed. 2021;60(18):10423-10429. doi:10.1002/anie.202016961
  2. Wu W, Wang W, Qi L, et al. Screening of xanthine oxidase inhibitors by liquid crystal-based assay assisted with enzyme catalysis-induced aptamer release. Anal Chem. 2021;93(15):6151-6157. doi:10.1021/acs.analchem.0c05456
Online Inquiry

For research and industrial use only, not for personal medicinal use.

Services
Online Inquiry

For research and industrial use only, not for personal medicinal use.