Aptamer Characterization Analysis Service

Alpha Lifetech has been providing aptamer related services for many years, and currently can provide comprehensive aptamer development schemes such as aptamer synthesis, SELEX screening, high-throughput sequencing, and aptamer optimization and characterization. Based on the advantages of SELEX method for screening aptamers, Alpha Lifetech has expanded more methods for screening aptamers.

Introduction to Aptamer Characterization Analysis

Affinity verification

The optimized aptamer affinity was verified by relevant techniques of the by aptamer binding assay. Examples include Isothermal Titration Calorimetry (ITC), Flow Cytometry (FCM), and Surface Plasmon Resonance (SPR), Microfluidics, etc. Aptamer affinity is usually expressed by the dissociation constant (KD), a physical quantity used to quantify the degree of molecular dissociation in a reversible reaction. The smaller the KD value, the more stable the complex, that is, the stronger the affinity; Conversely, the larger the KD value, the more unstable the complex and the weaker the affinity. This step is the key to identifying the performance of the aptamer, ensuring the aptamer affinity maturation and can bind to the target molecule with high affinity and selectivity in practical applications.

Fig.1 Process of aptamer affinity maturation. Reference source: Kinghorn AB, Fraser LA, 2017.

Function verification

In addition to affinity verification, the function of the aptamer needs to be verified. This includes verifying the aptamer's stability, specificity, and interactions with other molecules in a particular environment. The results of functional verification will directly affect the value of aptamers in scientific research and industrial applications.

Specific Verification

Competitive experiment: The ability of the aptamer to bind to a specific target is evaluated in the presence of other similar targets. If the aptamer can specifically bind to the target molecule without interference from other molecules, it indicates that it has high specificity.

Cross reaction experiment: The aptamer is bound to a series of related or unrelated targets to see if it only binds to a specific target, so as to verify its specificity.

Stability verification

Nuclease degradation experiment: The aptamers were exposed to different concentrations of nucleases and their degradation was observed. By comparing the degree of degradation at different time points, the anti-nuclease degradation ability of the aptamer can be evaluated.

Temperature and time stability experiment: The aptamer was placed under different temperature and time conditions to observe its structural and functional stability. This helps determine optimal storage and use conditions for aptamers.

Biological activity identification

The appropriate method is usually selected according to the specific application goal of the nucleic acid aptamer.

(1)Molecular level testing: Using molecular biology techniques, such as gel electrophoresis, Western blot, etc., to detect the complexes formed after the aptamer combines with the target molecule, or detect the changes in the expression level of the target molecule caused by the aptamer.

(2)Cell level testing: By using cell culture technology, aptamers are incubated with target cells to observe biological changes such as cell morphology, proliferation and apoptosis, and evaluate the biological activity of aptamers.

(3)Animal model testing: In appropriate animal models, nucleic acid aptamers are given by injection or drug administration, and physiological indicators and pathological changes of animals are observed to evaluate the biological activity and safety of the aptamers in vivo.