Aptamer Characterization Services

Creative Biolabs, the indisputable global leader in innovative drug development and production, has amassed extensive experience in delivering one-stop aptamer characterization services centered on the principles of high quality and time-saving. Our skilled scientists provide complete aptamer characterization services from many perspectives, based on an advanced technology platform.

Aptamer and Aptamer Characterization

Aptamers, which are short single-stranded oligonucleotides or peptides, extracted in vitro from randomized libraries and have a high affinity for certain compounds, can be used as promising molecular tools, diagnostic agents, and therapeutics. Aptamer characterization is to characterize and evaluate the properties of the interest aptamers for the efficiency of therapeutic applications, involving structure determination, binding kinetics measurement, and specificity assays, etc.

Fig. 1 Application areas of aptamers¹.

Aptamer Characterization Services at Creative Biolabs

Tertiary structure analysis

Computational methods: Online software such as mfold and NUPACK employ the nearest-neighbor model to calculate the free energy of oligonucleotide structures based on base pairing to estimate the secondary structure of aptamers.
Circular dichroism (CD): An approach that can reveal structural information on aptamers based on their capacity to absorb circularly polarized light.
Nuclear magnetic resonance (NMR)spectroscopy: It uses the interaction of radio-frequency electromagnetic energy with the nuclei of atoms within the aptamer as well as the target in a strong magnetic field to determine the tertiary structure of aptamers in aqueous solution.
X-ray crystallography: It works by exposing a crystalline sample to X-rays and evaluating the spatial distribution and intensity of the diffracted rays to identify the three-dimensional structure of aptamers.

Binding affinity measurement

Based on binding-induced changes in target fluorescence: It measures differences in the fluorescence of a small-molecule ligand when it is bound to an aptamer compared to when it is free in solution to analyze aptamer affinity and specificity.
Fluorescence polarization: It works on the basis that receptor-ligand interaction reduces either binding partner's degrees of freedom, which can be identified by measuring the polarization of fluorescent light emitted by one of the species.

Thermodynamic profiling

Isothermal titration calorimetry (ITC): It may offer detailed aptamer KD values ranging from nM to M, along with ligand-binding stoichiometry, enthalpy, and entropy.
Surface plasmon resonance (SPR): It can characterize binding affinity and stoichiometry, kinetic metrics such as kon and koff for determining the binding parameters of aptamers.
Microscale thermophoresis (MST): It utilizes that bound aptamers or ligands have distinct - usually lower - diffusion rates in solution than their free counterparts to characterize the KD of aptamers for small molecules.

Competition assays

Bead-based binding assays: A biotinylated complementary strand immobilizes the aptamer on streptavidin-coated agarose beads. It is compatible with aptamers separated by library-immobilized SELEX.
Gold nanoparticle (AuNP)-based assays: It works that unmodified AuNPs can transform ligand-aptamer binding into a colorimetric signal in order to screen the binding of aptamers to specific molecules.
Strand-displacement assays: It works by targeting a complementary DNA strand that has been hybridized with a part of the aptamer.

Importance of Aptamer Characterization

Understanding Aptamer Properties

Characterization provides valuable insights into the aptamer's properties, such as binding affinity, specificity, and selectivity. These properties determine the aptamer's effectiveness in its intended application, ensuring optimal performance.

Quality Control

Characterization ensures the quality and consistency of aptamers. It helps verify that the aptamers meet predefined specifications and fulfill required performance criteria. Quality control measures prevent batch-to-batch variations and ensure reliable results in research, diagnostics, or therapeutic applications.

Optimization of Aptamer Design

Safety Assessment

Aptamer characterization contributes to the safety assessment of aptamers used in clinical or diagnostic settings. Evaluating off-target binding and potential toxic effects is critical to minimize adverse reactions and ensure patient safety. Comprehensive characterization helps identify any potential risks associated with aptamer usage.

Creative Biolabs has a wide range of innovative chemical and biological technologies to meet the needs of customers in the rapidly expanding therapeutic aptamer research field. Please directly contact us for more details.

Reference

Li, Q, et al. "Aptamers: a novel targeted theranostic platform for pancreatic ductal adenocarcinoma." Radiation oncology (London, England) vol. 15,1 189. 5 Aug. 2020.

Questions & Answer

A: Aptamer specificity is typically determined by assessing its binding affinity to the target molecule compared to other non-target molecules. This can be done through competition assays or by measuring the binding constants using techniques like SPR.

A: Aptamer stability characterization helps determine the robustness and durability of aptamers under various conditions, such as temperature, pH, and storage conditions. It is crucial to assess stability for reliable and consistent performance. Aptamer stability can be assessed by subjecting the aptamers to different conditions and evaluating their binding activity over time. Techniques such as thermal denaturation studies and stability assays can be used to measure the stability of aptamers.

A: The key parameters assessed during aptamer characterization include binding affinity, specificity, stability, folding, kinetics, and any potential off-target interactions. These parameters collectively provide a comprehensive understanding of the aptamer's behavior.

For Research Use Only.