Fluorescence Assays for DNA Topoisomerase
Top Biosciences offers three proprietary, fluorescence-based assays specifically engineered for the discovery and identification of DNA topoisomerase inhibitors. As the only fluorescence-only assays in this field, our technology is protected by multiple US patents and provides a streamlined alternative to traditional methods.
Why Choose Our Assays?
Our platforms are designed with medicinal chemists and drug discovery labs in mind. By eliminating the need for biochemistry-heavy protocols, we enable rapid screening without specialized training.
No Gels Required: Completely eliminates the need for time-consuming agarose or polyacrylamide gel electrophoresis.
HTS-Ready: Fully optimized for High-Throughput Screening (HTS) to accelerate lead compound identification.
User-Friendly: Ideal for labs without extensive biochemistry facilities or specialized personnel.
Our Assay Suite
We provide three distinct screening tools to target different inhibitory mechanisms:
Catalytic Inhibitor Assays (Two Assays): Designed to identify compounds that interfere with the catalytic cycle of DNA topoisomerases.
Bacterial DNA Gyrase Poison Assay: A specialized HTS assay for identifying "poisons" that stabilize the covalent enzyme-DNA complex, a proven strategy for antibacterial drug development.
Contact Us
Advance your drug discovery program with our patented technology. For inquiries regarding our HTS assays and licensing, please contact our team:
Email: [email protected]
Products
1. Bacterial DNA Gyrase Fluorescence Assay Kit.
2. DNA Topoisomerase Fluorescence Assay Kit.
Top Biosciences offers three proprietary, fluorescence-based assays specifically engineered for the discovery and identification of DNA topoisomerase inhibitors. As the only fluorescence-only assays in this field, our technology is protected by multiple US patents and provides a streamlined alternative to traditional methods.
Why Choose Our Assays?
Our platforms are designed with medicinal chemists and drug discovery labs in mind. By eliminating the need for biochemistry-heavy protocols, we enable rapid screening without specialized training.
No Gels Required: Completely eliminates the need for time-consuming agarose or polyacrylamide gel electrophoresis.
HTS-Ready: Fully optimized for High-Throughput Screening (HTS) to accelerate lead compound identification.
User-Friendly: Ideal for labs without extensive biochemistry facilities or specialized personnel.
Our Assay Suite
We provide three distinct screening tools to target different inhibitory mechanisms:
Catalytic Inhibitor Assays (Two Assays): Designed to identify compounds that interfere with the catalytic cycle of DNA topoisomerases.
Bacterial DNA Gyrase Poison Assay: A specialized HTS assay for identifying "poisons" that stabilize the covalent enzyme-DNA complex, a proven strategy for antibacterial drug development.
Contact Us
Advance your drug discovery program with our patented technology. For inquiries regarding our HTS assays and licensing, please contact our team:
Email: [email protected]
Products
1. Bacterial DNA Gyrase Fluorescence Assay Kit.
2. DNA Topoisomerase Fluorescence Assay Kit.
T5 Exonuclease AT-Hairpin Fluorescence Assay
A Cost-Effective HTS Platform for Catalytic Inhibitor Discovery
This patented assay leverages the unique activity of T5 exonuclease on the (-) supercoiled plasmid pAB1, which carries a specific AT-hairpin structure. While T5 exonuclease completely degrades the supercoiled form, it cannot digest the relaxed form. This biochemical distinction allows for the rapid, fluorescence-based identification of inhibitors targeting various DNA topoisomerases.
Key Features & Validation
Intellectual Property & Publication
This technology is peer-reviewed and protected by a US patent:
Products
1. Bacterial DNA Gyrase Fluorescence Assay Kit.
2. DNA Topoisomerase Fluorescence Assay Kit.
This patented assay leverages the unique activity of T5 exonuclease on the (-) supercoiled plasmid pAB1, which carries a specific AT-hairpin structure. While T5 exonuclease completely degrades the supercoiled form, it cannot digest the relaxed form. This biochemical distinction allows for the rapid, fluorescence-based identification of inhibitors targeting various DNA topoisomerases.
Key Features & Validation
- Versatile Screening: Optimized to identify inhibitors for diverse DNA topoisomerases, T5 exonucleases, and DNA intercalators.
- HTS-Ready: Fully transitioned to a high-throughput format and validated with a Z'-factor > 0.5, ensuring high reproducibility for large-scale library screening.
- Proven Reliability: The assay's performance was successfully validated using a 50-compound library.
Intellectual Property & Publication
This technology is peer-reviewed and protected by a US patent:
- Publication: Deng Z, Leng F. A T5 Exonuclease-Based Assay for DNA Topoisomerases and DNA Intercalators. ACS Omega. 2021;6(18):12205-12212.
- Patent: Leng, F. and Deng, Z. T5 exonuclease-based method to identify DNA topoisomerase inhibitors. US Patent 11,732,286 B2, issued August 22, 2023.
Products
1. Bacterial DNA Gyrase Fluorescence Assay Kit.
2. DNA Topoisomerase Fluorescence Assay Kit.
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Supercoiling-Dependent Fluorescence Quenching (SDFQ) Assay
Ultra-High-Throughput Screening (uHTS) for Novel Topoisomerase Inhibitors
We have developed a proprietary method to produce fluorescently labeled circular DNA molecules (e.g., pAB1_FL905) at milligram scale. These molecules serve as powerful sensors for DNA topology. Because the assay requires only nanogram quantities of DNA per well, it is uniquely suited for ultra-High-Throughput Screening (uHTS) in 384-well or 1536-well formats.
Key Features & Validation
Intellectual Property & Publications
This uHTS technology is validated by high-impact research and protected by a US patent:
We have developed a proprietary method to produce fluorescently labeled circular DNA molecules (e.g., pAB1_FL905) at milligram scale. These molecules serve as powerful sensors for DNA topology. Because the assay requires only nanogram quantities of DNA per well, it is uniquely suited for ultra-High-Throughput Screening (uHTS) in 384-well or 1536-well formats.
Key Features & Validation
- Broad Applicability: Designed to identify inhibitors targeting bacterial DNA gyrase as well as human DNA Topoisomerases I and II.
- Validated Performance: Using this platform to screen the LOPAC and MLSMR libraries, we identified 155 novel inhibitors, including both catalytic inhibitors and gyrase poisons.
- Antibiotic Discovery: The structurally diversified scaffolds identified via this assay provide a robust foundation for developing next-generation antibiotics against multi-drug resistant pathogens.
Intellectual Property & Publications
This uHTS technology is validated by high-impact research and protected by a US patent:
- Publication 1: Gu M, et al. Fluorescently labeled circular DNA molecules for DNA topology and topoisomerases. Sci Rep. 2016;6:36006.
- Publication 2: Alfonso EE, et al. Novel and Structurally Diversified Bacterial DNA Gyrase Inhibitors... ACS Pharmacol Transl Sci. 2022;5(10):932-944.
- Patent: Leng, F. Labeled circular DNA molecules for analysis of DNA topology, topoisomerases, and drug screening. US Patent 9,890,414 B2, issued February 13, 2018.
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A High-Throughput Fluorescent DNA Cleavage Assay for Discovering Bacterial
DNA Gyrase Poisons
A High-Sensitivity HTS Platform for Discovering Bacterial DNA Gyrase Poisons
We have established a novel, fluorescence-based assay specifically designed to identify bacterial DNA gyrase poisons. The method leverages the stabilization of multiple gyrase-DNA cleavage complexes on a single plasmid. These complexes are trapped and then digested by Proteinase K, resulting in DNA fragmentation.
The unique strength of this assay lies in the use of T5 exonuclease, which selectively degrades the resulting linear and nicked fragments. This process "amplifies" the fluorescence signal loss (after SYBR Green staining), creating a highly sensitive readout for compound activity.
Key Features & Validation
Intellectual Property & Publication
This technology is documented in a high-impact preprint and protected by a recently issued US patent:
We have established a novel, fluorescence-based assay specifically designed to identify bacterial DNA gyrase poisons. The method leverages the stabilization of multiple gyrase-DNA cleavage complexes on a single plasmid. These complexes are trapped and then digested by Proteinase K, resulting in DNA fragmentation.
The unique strength of this assay lies in the use of T5 exonuclease, which selectively degrades the resulting linear and nicked fragments. This process "amplifies" the fluorescence signal loss (after SYBR Green staining), creating a highly sensitive readout for compound activity.
Key Features & Validation
- Mechanism-Specific: Precisely targets gyrase poisons that stabilize covalent enzyme-DNA complexes.
- Validated via Large-Scale Screening: The platform was successfully validated through pilot screens of the LOPAC1280, Maybridge (14k compounds), and 6K libraries.
- Commercial Readiness: All screens demonstrated a $Z'$-factor > 0.5, confirming the assay is robust and ready for large-scale commercial compound library screening.
Intellectual Property & Publication
This technology is documented in a high-impact preprint and protected by a recently issued US patent:
- Publication: Dias M, et al. A Fluorescence-Based, T5 Exonuclease-Amplified DNA Cleavage Assay for Discovering Bacterial DNA Gyrase Poisons. bioRxiv [Preprint]. 2023.
- Patent: Leng, F. and Dias, M. High throughput screening assay to identify DNA topoisomerase inhibitors. US Patent 12,234,504 B1, issued February 25, 2025.
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