Aptamer Screening through SELEX and Phage Display

Aptamers are single-stranded DNA or RNA sequences that fold into three-dimensional structures capable of recognizing a target of interest with affinity and specificity. The process for selecting a synthetic aptamer is known as in vitro selection or Systematic Evolution of Ligands by Exponential Enrichment (SELEX).

Aptamers are particularly advantageous as they are small in size, highly modifiable, stable at ambient temperatures, relatively simple to produce with limited batch-to-batch variation and have a low degree of immunogenicity. These desirable qualities enable the application of aptamers in drug delivery systems, therapeutics, diagnostics, biosensors and molecular imaging. Development of better aptamers with extremely high affinity and unparalleled specificity is vital.

Two Kinds of Aptamers

1) Nucleic acid-based aptamers

Nucleic acid-based aptamers (DNA aptamers and RNA aptamers or modified XNA aptamers) are small, highly structured DNA/RNA molecules, isolated from combinatorial libraries by SELEX. They have affinities for their target molecules similar to how antibodies work.

2) Peptide aptamers

Peptide aptamers are small combinatorial peptides that are selected to bind to specific sites on their target molecules, consisting of short, 5-20 amino acid residues long sequences, typically embedded as a loop within a stable protein scaffold. Peptide aptamers were isolated from combinatorial display libraries, such as phage display library, by various display techniques, such as phage display biopanning.

Development of better aptamers Platforms

In order to achieve this goal, Fusion BioLabs have made disruptive improvements in three key aspects: aptamer library design strategies (classic aptamer library vs structured library), various SELEX methodologies (beads-based, cell-SELEX, in vivo SELEX), and new chemical modifications. We help push aptamer research and applications to the next level.

1. Aptamer Library Strategies

• Standard Aptamer Library

Since SELEX technologies was developed in 1990, selection experiments begin with the chemical synthesis of a nucleic acid library containing a region of randomized nucleotides flanked by fixed primer binding regions. The standard random library approach can only cover the sequence space of libraries containing up to 25 randomized positions in theory. As the length of the random domain increases, the sequence space coverage of a random library drops off significantly, making it less likely to isolate the most active variants. However, many different sequences can fold into similar secondary structures that exhibit comparable activities. As long as their structured regions maintain stable base-pairing, the most active primary sequence can tolerate a high degree of variation. Thus, it is more plausible that the initial pool will contain less optimal versions of the strongest aptamer family, which can then be improved upon via reselection.

For nucleic acid aptamer development (DNA aptamer, RNA aptamer or XNA aptamer), Fusion BioLabs provide classic N40 Aptamer Library (40-uniformly randomized nucleotide domain) or N62 Aptamer Library (62-uniformly randomized nucleotide domain) for SELEX screening different targets.

For peptide aptamer development, Fusion BioLabs provide either cyclized C8C peptide phage library (8-amino acid cyclized peptide domain) or linearized X8 peptide phage library (8-amino acid linearized peptide domain), X16 peptide phage library (16-amino acid linearized peptide domain) and X24 peptide phage library (24-amino acid linearized peptide domain) for phage panning different targets.

• Structured Aptamer Library

Fusion BioLabs use rational library design strategy to increase the presence of functional sequences (aptamer candidates) in nucleic acid pools or peptide pools. For instance, certain secondary structure motifs can be integrated directly into the initial library to bias a selection towards more complex folds. Structurally important regions can be held constant in the starting library while other nucleotides or amino acids, such as those implicated in target binding, are randomized. This approach not only decreases the labor work but also enhances aptamer affinity and specificity of aptamer candidates

For nucleic acid aptamer development (DNA aptamer, RNA aptamer or XNA aptamer), Fusion BioLabs provide G-quadruplex Structured Aptamer Library (G-Quadruplex Motifs as Structured Library Elements), Hairpin Structured Aptamer Library (stem loop motifs as structured library elements) and Junctions Structured Aptamer Library (Junction Motifs as Structured Library Elements) for SELEX screening different types of targets.

For peptide aptamer development, Fusion BioLabs provide Affibody Phage Display Library, a structural scaffold library for phage panning different types of targets.

2. SELEX methodologies

Fusion BioLabs apply various SELEX methodologies to screening different types of your targets:

• Beads-based SELEX for screening targets with functional conjugate groups, such as peptides, native proteins or tagged proteins, et al.
• Capture-SELEX for screening small molecules and large molecules.
• Cell-based SELEX for screening targets located on the cell membrane (surfaced bound aptamer development) or inside the cell (internalization aptamer development).
• In vivo SELEX for select aptamers against living organisms.

 

3. Chemical modifications

Fusion BioLabs have successfully developed 2’-fluoro RNA aptamer, 2’-OMe RNA aptamer and L-RNA aptamer for some specific targets under special conditions.

 

Services we provided

Fusion Biolabs provide a broad range of unique aptamer screening services for custom aptamer development assays.

For detailed pricing information, please fill the form below for a custom quotation!