Protein Secondary & Tertiary Structure Analysis
Determine α-helix, β-sheet, and tertiary folding using advanced deconvolution algorithms.
Circular Dichroism Analysis
Wondering how stable your protein is, or whether your design folds as expected? Our Circular Dichroism analysis gives you fast, accurate answers—so you can move forward with confidence.
- Comprehensive Insights – Analyze secondary & tertiary structure in solution.
- Minimal Sample Requirement – Accurate results with only 50–100 μg.
- Thermal & Chemical Stability Testing – Tm determination with ±0.5 °C precision.
- Rapid, High-Resolution Data – Full spectra in minutes, 190–800 nm range.
- Validated Interpretation – Advanced algorithms ensure ±5% structural accuracy.
What Is Circular Dichroism?
Circular Dichroism (CD) is a spectroscopic technique that measures the differential absorption of left- and right-handed circularly polarized light by chiral molecules. This method is highly sensitive to the secondary and tertiary structures of biomolecules, especially proteins, peptides, nucleic acids, and complex biomolecular assemblies. By detecting subtle conformational differences, CD enables researchers to assess molecular folding, stability, and structural transitions under various conditions.
Typical Scientific Questions Circular Dichroism Can Answer
- What proportion of α-helix, β-sheet, and random coil structures does a protein exhibit?
- How does a protein unfold under increasing temperature or denaturant concentration?
- What is the refolding pathway after stress or ligand binding?
- Does a small molecule or metal ion induce structural rearrangements?
- Are biosimilars or engineered proteins structurally consistent with reference molecules?
Advantages of Our Circular Dichroism Service
Ultra-Low Sample Requirement — Efficient and Cost-Saving
Achieve accurate secondary structure analysis with as little as 50–100 μg of protein, minimizing material consumption for valuable samples.
High-Resolution Structural Insights — From Far-UV to Visible Range
Capture secondary, tertiary, and chromophore-related conformations with full-spectrum coverage (190–800 nm).
Thermal & Chemical Stability Profiling — Precision at Every Degree
Determine melting temperature (Tm) with ±0.5 °C accuracy and monitor unfolding pathways under controlled conditions.
Rapid Turnaround Without Compromising Quality
Generate complete spectra in less than 10 minutes, enabling high-throughput assessments for multiple candidates.
Validated Algorithms for Reliable Interpretation
Employ advanced deconvolution models (CDSSTR, SELCON3) to deliver quantitative secondary structure content with ±5% accuracy
Technical Services
Service Scope
Workflow and Instrumentation
Application
Sample Requirement
Deliverables
FAQ
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Scope of Circular Dichroism Services at Creative Proteomics
At Creative Proteomics, our Circular Dichroism (CD) services are designed to solve the structural and stability questions that matter most to your research and development projects. Whether you are characterizing a new protein candidate, validating biosimilarity, or ensuring formulation robustness, we provide data-driven insights that accelerate decision-making.
Thermal & Chemical Denaturation Studies
Profile unfolding curves and Tm with ±0.5 °C precision under thermal or chemical stress.
Folding and Refolding Kinetics
Monitor structural transitions in real-time, including intermediate states.
Advanced Option: Stopped-flow CD for millisecond-scale reaction kinetics and transient state detection.
Ligand and Cofactor Interaction Analysis
Detect conformational changes upon small molecule, metal ion, or excipient binding, supporting SAR studies.
Chiral Compound Stereochemistry
Assess optical activity and absolute configuration of chiral small molecules and macromolecules, ensuring accurate stereochemical analysis.
Batch Comparability & Biosimilarity Assessment
Confirm higher-order structure consistency between reference and biosimilar products.
Formulation Stability Assessment
Evaluate structural integrity under formulation conditions, including pH variation, ionic strength changes, and the presence of surfactants or stabilizers.
Nucleic Acid Conformation Studies
Characterize DNA/RNA forms (B-DNA, Z-DNA, G-quadruplex) for oligonucleotide and aptamer research.
Interested in RNA-focused projects? Learn more about our RNA Circular Dichroism Assay for G-quadruplex and secondary structure characterization.
Our Circular Dichroism Analysis Workflow
1
Sample Receipt & Quality Check
Verify purity (>90%), buffer compatibility (low UV absorbance), and concentration (0.1–1.0 mg/mL). Samples filtered (0.22 μm) and degassed prior to analysis.
2
Experimental Design
Select appropriate scanning range:
- Far-UV (190–260 nm) for secondary structure
- Near-UV (260–320 nm) for tertiary structure
- Full-range (180–800 nm) for chromophores
Temperature-controlled experiments (4–95 °C, 1 °C/min) for stability profiling.
3
Spectral Acquisition
Performed on Jasco J-1500 and Chirascan™ systems with high sensitivity (0.1 nm step size). Baseline correction and calibration ensure accurate spectra.
4
Data Analysis & Structural Interpretation
Secondary structure deconvolution, thermal stability modeling, and comparative studies.
5
Report Delivery
Comprehensive CD report including raw spectra, structural estimates, and interpretative discussion.
Circular Dichroisms Instrumentation & Technical Capabilities
Core Instrumentation
- High-Performance CD Platforms: Jasco J-1500 and Applied Photophysics Chirascan™ for accurate optical activity measurements.
- Temperature Control: 4 °C to 95 °C with precise thermal ramping for stability studies.
- Broad Wavelength Range: 180–800 nm, covering far-UV, near-UV, and visible regions.
Analytical Features
- Flexible Pathlengths: 0.01–1 cm for varying sample concentrations.
- Low Sample Volume: As little as 200 μL, ideal for limited or high-value samples.
- Advanced Deconvolution: Algorithms (CDSSTR, SELCON3) ensure secondary structure estimates with ±5% accuracy.
Jasco J-1500
Application Scenarios
Structural Integrity Verification for Protein Therapeutics
Used in biosimilar comparability studies and lot-to-lot consistency checks during biologics development to confirm folding and higher-order structure compliance with regulatory standards.
Pre-Formulation Screening for Biologics
CD supports excipient selection and buffer optimization by monitoring protein stability under different formulation conditions, minimizing aggregation risk before scaling up.
Protein Engineering and Variant Assessment
In structural biology and protein design projects, CD helps researchers validate engineered mutants for desired structural features or conformational flexibility, ensuring rational design success.
Characterization of Folding Pathways in Basic Research
Time-resolved CD is used in academic studies of folding kinetics, providing insight into intermediate states and misfolding mechanisms—essential for understanding protein folding diseases.
Nucleic Acid Research in Drug Discovery
CD identifies conformational changes in DNA/RNA, such as G-quadruplex formation in oncogene promoters or aptamer folding—key for oligonucleotide drug development.
Metal Ion and Ligand Interaction Mechanism Studies
Pharmaceutical and structural biology projects leverage CD to map conformational shifts upon ligand binding, supporting structure-activity relationship (SAR) analysis in early-stage drug discovery.
Stress Testing in Stability-Indicating Studies
Applied during forced degradation studies to understand thermal or chemical-induced unfolding, ensuring robustness of therapeutic proteins for regulatory submissions.
Sample Requirements for Circular Dichroisms Projects
| Parameter | Requirement |
| Sample Type | Purified protein, peptide, or nucleic acid |
| Minimum Quantity | 50–100 μg (typical for proteins) |
| Concentration | 0.1–1.0 mg/mL (optimal range for most CD measurements) |
| Buffer Conditions | Low UV absorbance; avoid high salt, detergents, aromatic additives |
| Preferred Buffers | 10 mM phosphate or sodium fluoride |
| pH Range | 6.0–8.0 |
| Volume Required | ≥ 200 μL (standard cell with 0.1 cm path length) |
| Purity | ≥90% by SDS-PAGE or SEC |
| Temperature | Store/transport at 4 °C; avoid freeze-thaw cycles |
| Avoid Additives | DTT, β-mercaptoethanol, high glycerol (>5%), high salt (>50 mM) |
| Additional Notes | Degas and filter (0.22 μm) before submission; frozen samples accepted upon request |
Deliverables: What You'll Receive
High-Quality Raw Data
Full CD spectra (Far-UV, Near-UV, or visible range), baseline-corrected and instrument-calibrated.
Processed & Interpreted Results
- Secondary structure estimation with α-helix, β-sheet, and random coil percentages.
- Comparative analysis for ligand interaction, batch comparability, or formulation studies.
- Thermal and chemical denaturation profiles with Tm values and transition curves.
Compliance-Ready Report
A detailed report in PDF format, including:
- Experimental conditions and parameters.
- Data quality validation and calibration logs.
- Comprehensive interpretation and structural discussion.
Optional Bioinformatics Analysis (upon request)
Advanced deconvolution (CDSSTR, SELCON3), folding/unfolding kinetic modeling, and data integration with structural predictions.
Far-UV CD Spectrum
Far-UV CD spectrum (190–260 nm) comparing native and denatured protein conformations, highlighting α-helix structural transitions.
Near-UV CD Spectrum
Near-UV CD spectrum (260–320 nm) illustrating aromatic side-chain and tertiary structural differences between native and denatured proteins.
Secondary Structure Deconvolution Charts
Bar chart showing α-helix, β-sheet, and random coil percentages before and after denaturation.
Thermal Denaturation Curve
Thermal unfolding curve with Tm (≈62 °C) determined from ellipticity changes. Sigmoidal transition indicates protein melting temperature and structural stability under heat stress.
Ligand Interaction Difference Spectrum
Difference CD spectrum highlighting structural changes upon ligand binding. Positive and negative peaks indicate local conformational adjustments in secondary structure regions.
Folding/Refolding Kinetic Profiles
Time-resolved CD profiles for folding and refolding processes. Kinetic curves reveal intermediate states and folding rates critical for protein stability assessment.
Circular Dichroism vs Other Structural Analysis Techniques: Which Should You Choose?
| Feature | Circular Dichroism (CD) | FTIR | Fluorescence | NMR | X-ray Crystallography | DSC |
| Structural Insight | Secondary & Tertiary Structure | Secondary Structure | Tertiary Structure (local) | Atomic-level 3D Structure | Atomic-level 3D Structure | No Structure (Thermal Only) |
| Sample State | Solution | Solution | Solution | Solution | Crystal | Solution |
| Sample Amount | 50–100 μg | ~100 μg | <50 μg | >5 mg | mg-level | >0.5 mg |
| Analysis Speed | Minutes | Minutes | Minutes | Days–Weeks | Weeks | Hours |
| Quantitative Accuracy | ±5% for Secondary Structure | Moderate | Low | High | High | N/A |
| Thermal Stability Testing | Yes (with structural correlation) | Limited | Yes (qualitative) | No | No | Yes (detailed thermogram) |
| Cost Level | Moderate | Low | Low | High | High | Moderate |
| Data Interpretation | Easy | Moderate | Easy | Complex | Complex | Moderate |
| Best Use Case | Folding & Stability Screening | Buffer-Insensitive Conditions | Detecting Local Tertiary Changes | High-Resolution Structural Study | Binding Site Mapping for Drug Design | Thermal Stability Only |
You May Want to Know
What types of molecules can be analyzed by Circular Dichroism?
CD is suitable for chiral biomolecules such as proteins, peptides, nucleic acids, and some carbohydrates. It can also detect conformational changes in complexes like protein-ligand assemblies.
How accurate is CD in determining secondary structure content?
CD provides highly reliable relative estimates for α-helix, β-sheet, and random coil structures. With validated deconvolution algorithms (e.g., CDSSTR, SELCON3), the accuracy is typically within ±5%.
What wavelength regions are used, and what information do they provide?
Far-UV (190–250 nm): Secondary structure (α-helix, β-sheet, random coil).
Near-UV (250–320 nm): Tertiary structure and aromatic side-chain environment.
Visible region: Cofactor or chromophore binding studies.
Can CD detect minor structural changes caused by formulation or buffer changes?
Yes. CD is highly sensitive to subtle conformational alterations, making it ideal for monitoring structural integrity under varying pH, ionic strength, or excipient conditions.
Is CD compatible with all buffers?
Buffers with high UV absorbance (e.g., Tris, imidazole) should be avoided. Low-UV absorbing buffers like phosphate are preferred.
Can CD be used to study protein folding kinetics?
Yes. Time-resolved CD enables real-time observation of folding and refolding processes, allowing analysis of intermediate states and kinetic parameters.
Is CD compatible with membrane proteins or proteins in detergents?
CD can analyze membrane proteins; however, detergents and high UV-absorbing components may interfere with measurements. Low-absorbance detergents and buffer optimization are recommended.
What is the minimum pathlength required for low-concentration samples?
We offer flexible pathlengths down to 0.01 cm, allowing accurate measurement even for low-volume or dilute samples.
Does CD provide absolute structure information like X-ray or NMR?
No. CD offers secondary and tertiary structural estimation in solution but does not deliver atomic-resolution structures. For high-resolution details, CD is complementary to X-ray or NMR studies.
Can CD detect nucleic acid conformational changes such as G-quadruplex formation?
Yes. CD is widely used to distinguish between DNA/RNA conformations such as B-DNA, Z-DNA, and G-quadruplex structures under different ionic conditions.
