What is an Aptamer? First, a quick reminder: Aptamers are short, single-stranded DNA or RNA oligonucleotides that bind to a specific target with high affinity and specificity. They are often called "chemical antibodies." The Core Service: SELEX (The Screening Process) The service revolves around executing a SELEX (Systematic Evolution of Ligands by EXponential enrichment) campaign. This is an iterative, in-vitro combinatorial chemistry process that screens a vast random library (10^14 - 10^15 unique sequences) to find the few that bind your target. A standard SELEX workflow includes: Library Design & Synthesis: Creating the initial random oligonucleotide pool. Incubation: The library is exposed to the target. Partitioning: Bound sequences are separated from unbound ones (the most critical step, varying by target type). Amplification: The bound sequences are amplified (usually by PCR for DNA, RT-PCR for RNA). Counter-Selection (Negative Selection): To increase specificity, the pool is exposed to non-target surfaces (e.g., immobilization matrix, related proteins) to remove non-specific binders. Repetition: Steps 2-5 are repeated for 8-15 rounds until a high-affinity pool is enriched. Cloning & Sequencing: The final pool is cloned, and individual aptamer sequences are identified via Next-Generation Sequencing (NGS). Bioinformatics & Analysis: NGS data is analyzed to identify candidate sequences, often clustered into families based on sequence/structure motifs. Characterization: Top candidates…
Aptamer Capture-SELEX Service refers to a specialized, outsourced process where a company or academic core facility performs the entire Capture-SELEX procedure to develop DNA or RNA aptamers for a client's specific target molecule. This is a crucial service for researchers and companies who need high-affinity, specific aptamers but lack the specialized equipment, expertise, or time to run the SELEX process in-house. Let's break down what this service entails. 1. What is Capture-SELEX? First, understand the standard SELEX (Systematic Evolution of Ligands by EXponential enrichment). It's an iterative process to select aptamers from a vast random oligonucleotide library (10^14 - 10^15 different sequences). Capture-SELEX is a specific variant designed primarily for small molecules or targets that are difficult to immobilize directly on a solid support without affecting their structure/function. The Key Difference: Instead of immobilizing the target itself, a short, complementary "capture strand" is immobilized on beads or a surface. The initial ssDNA library is designed with a region complementary to this capture strand. The target is free in solution. How it Works: The library is bound to the surface via the capture strand. The target molecule is introduced in solution. Only library sequences that fold into a structure capable of binding the target will undergo a conformational change. This binding event often weakens or…
What is Live Cell SELEX? Traditional SELEX uses purified target proteins. Live Cell SELEX uses intact, living cells in their native state. This is crucial because: It selects for aptamers that bind to targets in their natural conformation and post-translational modifications (e.g., glycosylation). It inherently selects for cell-specificity (e.g., cancer cell vs. healthy cell) without needing to know the exact molecular target upfront. It can discover aptamers against unknown or membrane-bound targets that are difficult to purify. Core Workflow of a Typical Service A full-service provider will manage the entire pipeline: 1. Project Design & Consultation Target Cell Line Definition: Defining the "positive" cell line (e.g., patient-derived cancer cells, activated immune cells). Counter-Selection Strategy: Choosing the "negative" cell line(s) (e.g., healthy counterpart, isogenic control) to eliminate non-specific binders. Library Design: Recommending or customizing the starting random oligonucleotide library (length, modifications like 2'-F pyrimidines for RNA aptamers for stability). 2. The Selection Phase (The Iterative SELEX Cycles) Incubation: The random library is incubated with the counter-selection cells. Unbound/non-specific sequences are collected. Positive Selection: The pre-cleared library is incubated with the target cells. Cells are washed stringently. Recovery: Cell-bound aptamers are recovered (e.g., by cell lysis, heat elution, or protease treatment). Amplification: Recovered sequences are amplified by PCR (for DNA) or RT-PCR (for…
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 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. Access to All Binding Sites: All surfaces of the target are available for aptamer binding, increasing the diversity of potential aptamers discovered. 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. Better for Small Molecules and Peptides: Ideal for targets that are difficult to immobilize without blocking their functional groups. Mimics Physiological Conditions: More closely replicates how the aptamer will interact with its target in real-world applications…
What is Classical SELEX? SELEX is an iterative, in vitro selection process used to isolate single-stranded DNA or RNA molecules (aptamers) that bind with high affinity and specificity to a target (e.g., a protein, small molecule, cell, or virus). The "classical" method refers to the original, well-established protocol involving: Incubation: A vast, random-sequence nucleic acid library (10^14 - 10^15 different sequences) is exposed to the target. Partitioning: Unbound sequences are washed away; bound sequences are retained. Elution: The bound sequences are recovered. Amplification: The recovered sequences are amplified by PCR (for DNA) or RT-PCR (for RNA). Repetition: This cycle (typically 8-15 rounds) is repeated, enriching the pool for the strongest binders. Components of a Classical SELEX Service A full-service provider typically manages the entire pipeline: 1. Project Design & Consultation Target Characterization: Discussing the target's properties (purity, stability, availability). Selection Strategy: Deciding on immobilization method (e.g., target immobilized on beads, or "counter-SELEX" to eliminate binders to the immobilization matrix or similar non-target molecules). Library Design: Choosing DNA or RNA, length of the random region (typically 20-60 nt), and fixed primer regions. 2. The SELEX Process Execution Library Synthesis: Chemical synthesis of the initial random library. Cycle Management: Performing the repetitive rounds of binding, washing, elution, and amplification under optimized buffer and stringency…
The unique secondary and tertiary structures of aptamers provide the specificity to detect even small structural changes in the target molecule, including the presence or absence of methyl or hydroxyl groups or differences in enantiomeric configurations. Aptamers that bind specific targets are identified through a process known as Systematic Evolution of Ligands by Exponential enrichment (SELEX) in which binding molecules are selected from a large and diverse library of nucleic acids (either DNAs or RNAs). In this process, the nucleic acid library is incubated with the target molecule. Non-binding nucleic acids are then washed away, leaving behind only the molecules that have a capacity to bind to the target molecule. The nucleic acids that are not washed away are then used to create a new library of nucleic acids that is enriched for the subset that binds the desired target. Repeating this selection-cycle on each subsequent library with increasing stringency of binding (e.g. lower concentration of target), ensures that nucleic acids that bind to the target with both high specificity and high affinity are enriched. Aptamers are short, single-stranded oligonucleotides (DNA or RNA) that bind to specific target molecules with high affinity and specificity. They are often called "chemical antibodies."…
