Assay Development
& Screening
Biochemical and cell-based assays employing recombinant proteins, normal and engineered cell lines and reporter cell systems can be performed for a wide range of biological target classes. Reagents for assay development and screening can either be commercially purchased or produced at our reagent generation laboratories. Liquid handling systems, multimode plate readers, multiplex, phenotypic, label-free and radiometric screening platforms provide wide support for high-throughput screening campaigns.
Platform Technologies
- Luminescence
- TR-FRET/HTRF
- Fluorescence Polarization
- Absorbence
- Alpha Screen®
- High Content Imaging
- Flow Cytometry
- NanoBRETTH™
- qPCR
- SPR (Biacore)
- WES Protein Simple
- Radiometric (SPA, filter binding)
- Light Scattering
- Patch Clamp
- Next Generation Sequencing (NGS)
Models
- Recombinant Proteins
- Transient/Stable Cell Lines
- Reporter Cell Lines
- Mammalian Cell Lines
- PBMCs derived from Peripheral
- Primary Immune Cell Subtypes
- Patient Derived Primary Cells
- iPSCS; Tissue Homogenates Serum/Plasma
Therapeutic Areas
- Inflammation
- Oncology
- Immuno-Oncology
- Metabolic Diseases
- Fibrosis
- Autoimmune Disorders
Target Classes
- Enzymes; GPCR
- Ion Channels
- Transcription Factors
- Kinases
- Transporters
- Nuclear Hormone Receptors
Automation
- Biomek Fx/Nx Automated Workstation. with Cytomat
- Echo 550 Acoustic Liquid Handling System
Targeted Protein Degradation
Proteolysis targeting chimera or PROTAC is a paradigm shifting novel drug discovery strategy providing significant opportunity to transform the way medicinal chemistry and discovery programs are planned and executed. With sub-stoichiometric catalytic mode of action, PROTAC reduces the need for target occupancy as necessitated with traditional small molecule inhibitors and is being actively explored for Oncology and Inflammatory Disorders.
Key assays to support degrader programs need to take into consideration the key aspects of ubiquitin proteasomal machinery and E3 ligases involved to make hetero-bifunctional degraders or molecular glues.
- Binding assay: fluorescence polarization based competitive binding assay to evaluate binary binding affinity
- Ternary complex formation assay: AlphaScreen®, AlphaLISA®, or TR-FRET based assay to determine the formation of ternary complex
- Protein degradation assay: reporter complementation assays with customized stable cell lines to determine intracellular protein degradation in presence of degraders and DC50 and Dmax generation
Other Assays supporting the TPD Platform:
- Evaluating the ubiquitination of target proteins
- Western blot analysis of degradation of target protein
- Competition assays using unconjugated recruitment motifs
Agro Solutions
Aragen develops custom solutions to help farmers safeguard their harvest. As a partner to top agrochemical companies for over 10 years, we have wide experience in agrochemical research. Our scientists have been supporting screening assays in insecticidal, fungicidal and herbicidal site of action studies. We provide end-to-end support in compound screening and hit identification. Our team has developed a diversity of assays to identify the compound site of action in insects and plants, including mitochondrial, cell membrane, amino acid related, neurotransmitter and photosystems I and II.
Crop Protection Support
Screening for Insecticidal Site of Action (ISOA)
- Electron transport chain assay using mitochondria from insects and nematodes to confirm the site of action of complexes I, III and IV or II and III in respiratory disruptor classes
- Oxidative phosphorylation assay using Sf9 cells: a cell-based assay using Sf9 cells to confirm the site of action of mitochondria complex V in respiratory disruptor classes
- Acetylcholinesterase inhibition assay using insect source: an off-target assay to confirm neurotransmitter inhibition
- Mitochondrial toxicity: a cell-based assay to confirm respiratory disruptor classes
- Proliferation assay using Sf9 cells: a cell-based assay to confirm the mode of action involved in the control of insect growth and development
Screening for Fungicidal Site of Action (FSOA)
- Electron transport chain assay using mitochondria from fungi to confirm the site of action of complexes I, III and IV or II and III in respiratory disruptor classes
- Ergosterol biosynthesis inhibition (EBI) screen: a cell-based assay using various fungi to confirm the inhibition of biosynthesis pathway using absorbance and LCMS platform technology
Screening for Herbicidal Site of Action (HSOA)
- Acetolactate synthase enzymatic assay (ALS): a biochemical assay to confirm amino acid inhibitors
- Photosystem I & II assay using thylakoids prepared from leaf sample to confirm the herbicidal action in photosynthesis using OxygraphPlus
- Carotenoid pathway analysis (CRTB) using micro algae: a cell based assay to confirm the site of action of the carotenoid pathway LC/DAD
- HPPD assay: a complex assay to confirm the enzyme inhibition of 4-hydroxyphenylpyruvate dioxygenase (HPPD) inhibitors
- Protoporphyrinogen oxidase (PPO): an enzymatic assay to confirm the site of action of protoporphyrinogen oxidase (PPO) inhibitors
- THL assay: a simple biochemical assay to identify reactive compounds by assessing their reaction with glutathione (GSH), thereby eliminating the possibility for mammalian toxicity
GMO Testing & Crop Breeding Support
- Adventitious presence testing for BT cotton
- qPCR technique developed and tested for: DOW281-24-236, 3006-210-23, MON531, MON15985, MON88913
- Molecular marker analysis for crop breeding like SNPs
- Zygosity testing
- Copy number analysis
- Marker assisted selection
In Vitro Toxicology – Toxvit
We offer a broad suite of reliable toxicity testing solutions to enable go/no-go pipeline decisions:
- Modern toxicity testing technology
- Large scale operations with high-throughput equipment
- Reduced cost and testing times
- More robust mechanistic data
- Decisions on human relevant dose levels
- Validated testing protocols
In Vitro Toxicity Solutions
Multi-Parametric Cytotoxicity
- IC50 determination
- Cell loss/death (fluorescent imaging)
- Nuclear size and morphology (fluorescent imaging)
- Cell membrane permeability (fluorescent imaging)
- ~50 different cell lines of human origin available for cytotoxicity testing
Genetic Toxicity
- AMES MPF assay using Xenometrix AG-ANIARA microplate kit method (OECD 471)
- In vitro micronucleus assay using CHO, CHL, A549 cell lines and human PBMCs including cytotoxicity assessment on high content screening (HCS) platform (OECD 487)
- In vitro chromosomal aberration assay using CHO cell line and human PBMCs including cytotoxicity assessment (OECD 473)
- Mammalian cell gene mutation assay: MLA (L5178YTk+/- cell line) and HPRT (CHO-K1 cell line) mutations (OECD 490/476)
- In vitro comet assay using PBMC, CHO, CHL test systems on HCS platform
- H2AX double strand DNA damage response assay using A549, HepG2 and BEAS-2B cell lines on HCS platform
Dermal Toxicity
- In vitro irritation using 3D reconstructed human epidermis models (OECD 439)
- In vitro skin corrosion using 3D reconstructed human epidermis models (OECD 431)
- Phototoxicity assay using Balb/c-3T3 cells
- Skin sensitization assay using KeratinoSens cell line (OECD 442D)
- Skin sensitization by directing peptide reactivity assay (OECD 442C)
Ocular Toxicity
- Short term exposure assay using corneal epithelial cells (Statens Seruminstitut Rabbit Cornea SIRC cell line) (OECD 491)
- EpiOcular™ eye irritation assay using 3D reconstructed human corneal epithelium tissue model (OECD 492)
Hepatotoxicity Panel
- Using HepG2, HUH7, Hep3B cell lines and human iPSC HepRG spheroid cultures
- Drug-induced phospholipidosis and steatosis
- Lysosomal trapping (lysosomotropism)
- Cholestasis
- Mitochondrial permeability transition
Cardio Toxicity
- hERG safety screening using Invitrogen’s Predictor™ hERG fluorescence polarization assay kit
- 3D microtissue-based cardiotoxicity assay using human iPSC-CM (cardiomyocytes) spheroid cultures
Mitochondrial Toxicity
- Mitochondrial (Glu/Gal) assay in glucose and galactose supplemented media
- Mitochondrial membrane potential and cytochrome release
Neuro Toxicity
- Developmental neurotoxicity
- Neurite outgrowth
- Synaptogenesis assay
Endocrine Disruption Screening
- Estrogen receptor and androgen receptor binding assays using polar screen, human full-length ER (alpha/beta) competitor assay and polar green rat androgen receptor competitor assay
- Fluorescence polarization assay kits using ligand binding assays for ER and AR from rat membrane preparations (OPPTS/OCSPP 890.1250, OPPTS/OCSPP 890.1150)
- Steroidogenesis assay using H295R cell line with testosterone parameter assay kit and estradiol parameter assay kit (R&D systems) (OECD 456)
- Aromatase assay (OPPTS/OCSPP 890.1200)
Toxicogenomic Profiling
- Toxicological gene regulation studies through relative and quantitative expression of metabolic and toxicological pathway regulatory biomarkers using RT PCR and NGS platforms
Toxicity Testing Platforms
- High content imaging (thermo scientific cell insight) CX7 platform
- Multimode reader: Perkin Elmer Envision 2104 platform
- Flow cytometry: BD FACS verse platform
- Agilent platform for HPLC
- RT PCR-Applied Biosystems QuantStudio 6Flex platform
- Next generation sequencing (NGS) Illumina platform
Toxicity Testing Technologies
- Primary cell culture technology
- Pluripotent stem cell technology (PSC)
- 3D microtissue culture technology
