Solution-phase SELEX Service for Aptamers

What is Solution-Phase SELEX?

SELEX (Systematic Evolution of Ligands by EXponential Enrichment) is the iterative process used to discover aptamers—single-stranded DNA or RNA molecules that bind to a specific target with high affinity and specificity.

Solution-Phase SELEX refers to performing the selection process with the target molecule free in solution, rather than immobilized on a solid surface (like beads or a column). This often involves a partitioning step that separates bound from unbound sequences using a method like filtration, capillary electrophoresis, or magnetic bead capture of the target.

Key Advantages of Solution-Phase SELEX

1. Preservation of Native Target Conformation: The target is in its natural, free state in solution. This is crucial for complex targets like membrane proteins, which can denature or present epitopes unnaturally when immobilized.

2. Access to All Binding Sites: All surfaces of the target are available for aptamer binding, increasing the diversity of potential aptamers discovered.

3. Avoidance of Non-Specific Binding to Solid Support: Reduces background noise from library sequences sticking to the immobilization matrix (e.g., sepharose beads, plastic wells), leading to cleaner selections.

4. Better for Small Molecules and Peptides: Ideal for targets that are difficult to immobilize without blocking their functional groups.

5. Mimics Physiological Conditions: More closely replicates how the aptamer will interact with its target in real-world applications (e.g., diagnostics, therapeutics).

Typical Solution-Phase SELEX Workflow (as offered by a service)

A professional service will manage this entire, labor-intensive process:

1. Project Consultation & Design:

   • Target Discussion: Nature of the target (protein, small molecule, cell), purity, quantity, and buffer conditions.

   • Selection Strategy: Choosing the optimal solution-phase partitioning method (e.g., nitrocellulose filter binding, magnetic bead capture of tagged targets, capillary electrophoresis).

   • Library Design: Customizing the starting random library (length of random region, fixed primer sequences, modified nucleotides for nuclease resistance).

2. Selection Rounds (Cycles):

   • Incubation: The naïve oligonucleotide library is incubated with the target in solution under carefully optimized conditions (buffer, temperature, time).

   • Partitioning: The key step. Bound sequences are separated from unbound ones.

     • Filter Binding: Aptamer-target complexes are captured on a nitrocellulose filter; unbound DNA/RNA passes through.

     • Magnetic Bead Capture: If the target is biotinylated or has a tag, it can be captured with magnetic beads after complex formation.

     • CE-SELEX: Uses capillary electrophoresis to physically separate the bound complex (which migrates differently) from free library—extremely high stringency.

   • Elution & Recovery: Bound aptamers are dissociated from the target (e.g., by heat, denaturants) and collected.

   • Amplification: The recovered pool is amplified by PCR (for DNA) or RT-PCR (for RNA).

   • Purification: The amplified pool is purified for the next round.

   • Counter-Selection (Negative Selection): Often included to remove sequences that bind to common off-targets or the partitioning matrix itself (e.g., filters, beads without target).

3. Monitoring & Stringency Increase:

   • The enrichment of binding sequences is monitored (e.g., by qPCR, gel shift assays).

   • Selection conditions are made more stringent over rounds (e.g., reduced target concentration, increased wash stringency, added competitors) to select for the highest-affinity binders.

4. Sequencing & Bioinformatics:

   • The final enriched pool is cloned and sequenced via Next-Generation Sequencing (NGS).

   • Bioinformatics analysis identifies candidate families of aptamers based on sequence homology, folding, and frequency.

5. Characterization & Validation (Optional but critical):

   • Synthesis: Top candidate sequences are chemically synthesized.

   • Binding Analysis: Affinity (Kd) and specificity are measured using techniques like Surface Plasmon Resonance (SPR), Bio-Layer Interferometry (BLI), or Electrophoretic Mobility Shift Assay (EMSA).

   • Functional Assays: Testing aptamer performance in the intended application (e.g., inhibition of protein function, detection in a diagnostic format).

What to Look for in a Solution-Phase SELEX Service Provider

• Expertise & Proven Track Record: Look for experience with your target class (proteins, cells, small molecules) and publications/case studies.

• Flexibility in Partitioning Methods: Do they offer multiple techniques (filter binding, magnetic, CE-SELEX) to recommend the best for your target?

• NGS & Advanced Bioinformatics: Essential for deep analysis of the enriched pool. Avoid services that rely only on cloning and Sanger sequencing.

• Quality of Materials: Inquire about the libraries they use and their quality control processes.

• Transparent Process & Communication: Regular updates, clear timelines, and detailed final reports are crucial.

• Downstream Services: Can they provide full characterization (Kd, specificity), truncation, chemical modification (2’-F, 2’-O-Me), and large-scale production?

• Cost & Timeline: Typically ranges from $15,000 to $50,000+ and takes 3 to 6 months for the selection and initial sequencing, with additional time/cost for characterization.

Conclusion

Outsourcing to a solution-phase SELEX service is an excellent strategy to accelerate aptamer discovery without investing in the specialized expertise and equipment in-house. By choosing a provider with a robust solution-phase platform and strong analytical capabilities, you significantly increase the chances of obtaining high-quality, functional aptamers tailored for your specific application in therapeutics, diagnostics, or research.

When requesting a quote, be prepared to discuss: Your target’s identity, purity, available quantity, desired aptamer type (DNA/RNA/modified), and the intended final use.