SELEX Aptamer Screening

1. What is SELEX?

SELEX stands for Systematic Evolution of Ligands by EXponential enrichment.

It is a laboratory technique used to identify aptamers—short single-stranded DNA or RNA sequences that can bind specifically to a target molecule (proteins, small molecules, cells, or even viruses).

Aptamers act similarly to antibodies but are synthetic, highly stable, and can be chemically modified.

2. Principle of SELEX

The SELEX process is based on iterative rounds of selection and amplification:

1. Library Preparation

   • Start with a large randomized pool of oligonucleotides (typically 10^13–10^15 unique sequences).

   • Each sequence is a potential aptamer candidate.

2. Binding (Target Incubation)

   • Incubate the library with the target molecule.

   • Only sequences that can bind the target will stay attached; non-binders are washed away.

3. Partitioning (Separation of Binders and Non-binders)

   • Physically separate bound sequences from unbound sequences.

   • Techniques depend on the target (magnetic beads, affinity columns, etc.).

4. Elution

   • Bound sequences are eluted (released) from the target.

5. Amplification

   • The eluted sequences are amplified using PCR (for DNA aptamers) or RT-PCR (for RNA aptamers).

   • This generates an enriched pool for the next round.

6. Iterative Rounds

   • Steps 2–5 are repeated for 8–15 rounds to gradually enrich sequences with high affinity and specificity for the target.

7. Sequence Identification

   • After sufficient enrichment, sequences are sequenced (Sanger or NGS).

   • Top candidates are synthesized and tested for binding efficiency.

3. Variants of SELEX

• Cell-SELEX: Targets whole cells instead of purified proteins. Useful for biomarker discovery.

• In vivo SELEX: Selection occurs inside living organisms.

• Toggle SELEX: For broad-spectrum aptamers that recognize multiple related targets.

• High-Throughput SELEX: Uses NGS for faster and more comprehensive analysis.

4. Advantages of Aptamers via SELEX

• High specificity and affinity for the target (Kd in nM–pM range).

• Small size (20–80 nucleotides), can penetrate tissues easily.

• Chemically synthesized → no batch-to-batch variation.

• Stable at high temperatures; easy to modify chemically.

• Non-immunogenic (safe for therapeutic applications).

5. Applications of SELEX Aptamers

1. Diagnostics

   • Biosensors, ELISA alternatives, detection of biomarkers, viruses, or bacteria.

2. Therapeutics

   • Targeted drug delivery, inhibition of proteins or receptors (like Macugen for AMD treatment).

3. Research Tools

   • Protein purification, molecular recognition studies, imaging probes.

4. Environmental and Food Safety

   • Detect toxins, pathogens, or pollutants.

6. Workflow Summary

If you want, I can draw a clear diagram of the SELEX workflow that shows how the aptamer pool evolves and is enriched over rounds—it makes this process much easier to visualize.