An aptamer library is a diverse pool of nucleic acid sequences (DNA or RNA) from which specific aptamers—short oligonucleotides that bind to target molecules with high affinity—can be selected. Constructing a high-quality library is the foundation of aptamer screening technologies like SELEX. 2. Key Components of an Aptamer Library Randomized Region The central portion of the aptamer, typically 20–60 nucleotides, is randomized to generate diversity. Example: N20–N40 where N = A, T/U, G, or C. The diversity determines the probability of finding high-affinity binders. Flanking Constant Regions Short sequences (~15–25 nt) at both ends of the randomized region. Functions: Primer binding sites for PCR amplification. Stability and structural constraints. Overall Length Usually 40–100 nucleotides, balancing structural complexity and amplification efficiency. 3. Steps of Library Construction Design of Oligonucleotides Include random regions flanked by known primer sequences. Example structure:5'-[Primer]-N40-[Primer]-3' Chemical Synthesis Use solid-phase DNA/RNA synthesis to generate the oligonucleotides. Random nucleotides are incorporated using a controlled mixture of A, T/U, G, C. Amplification (for DNA libraries) PCR amplifies the synthesized sequences. RNA libraries require in vitro transcription from DNA templates. Purification Remove truncated or incomplete sequences. Methods: PAGE purification or HPLC. Quality Control Ensure correct length, diversity, and absence of biases.…
Customized Aptamer Selection refers to a tailored process of identifying and developing aptamers—short, single-stranded DNA or RNA molecules—that specifically bind to a target molecule (proteins, small molecules, cells, or pathogens) according to a client’s specific requirements. Unlike standard aptamer screening, it focuses on individualized targets, binding conditions, and functional needs. Key Features: Target Specificity: Aptamers are selected for high affinity and specificity to a particular target. Flexible Design: Can be designed for proteins, peptides, small molecules, ions, or whole cells. Binding Conditions Customization: pH, temperature, ionic strength, or buffer system can be tailored. Functional Application: Aptamers can be developed for diagnostics, therapeutics, biosensors, or research. High-Throughput & Efficiency: Advanced techniques allow rapid screening for optimal aptamers. Typical Workflow: Target Analysis: Understanding target structure and function. Library Preparation: Generate a diverse pool of oligonucleotides. SELEX (Systematic Evolution of Ligands by EXponential enrichment): Iterative selection process to enrich high-affinity aptamers. Binding Affinity Testing: Determine Kd (dissociation constant) and specificity. Sequence Optimization & Modification: Chemical modifications for stability or functionalization. Delivery of Customized Aptamer: Ready for research, diagnostics, or therapeutic use. Common Applications: Diagnostics: Biosensors for disease markers. Therapeutics: Targeted drug delivery. Research Tools: Protein purification or molecular imaging. Environmental Monitoring: Detection of…
“High-throughput aptamer screening” is a method used to rapidly identify aptamers—short single-stranded DNA or RNA molecules—that can bind specifically to a target molecule, such as a protein, small molecule, or even whole cells. Let’s break this down in detail: 1. What Are Aptamers? Aptamers are oligonucleotides (DNA or RNA) that fold into specific three-dimensional shapes allowing them to bind with high affinity and specificity to their targets. They function similarly to antibodies but are synthetic, smaller, more stable, and can be chemically modified. 2. High-Throughput Screening (HTS) in Aptamer Discovery Traditional aptamer discovery uses SELEX (Systematic Evolution of Ligands by Exponential Enrichment), which involves multiple iterative rounds of binding, separation, and amplification. High-throughput aptamer screening accelerates this process by using automation and large-scale technologies to simultaneously test thousands to millions of sequences against the target. 3. Key Techniques in High-Throughput Aptamer Screening Microarray-Based Screening Thousands of aptamer candidates are immobilized on a chip. The target (protein, small molecule, or cell) is fluorescently labeled and applied. Aptamers that bind the target emit signals detected by imaging. Next-Generation Sequencing (NGS)-Coupled SELEX After each SELEX round, sequences are analyzed via NGS. Sequence enrichment patterns reveal high-affinity aptamer candidates without the need for extensive…
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: Library Preparation Start with a large randomized pool of oligonucleotides (typically 10^13–10^15 unique sequences). Each sequence is a potential aptamer candidate. 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. Partitioning (Separation of Binders and Non-binders) Physically separate bound sequences from unbound sequences. Techniques depend on the target (magnetic beads, affinity columns, etc.). Elution Bound sequences are eluted (released) from the target. 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. Iterative Rounds Steps 2–5 are repeated for 8–15 rounds to gradually enrich sequences with high affinity and specificity for the target. Sequence Identification After…
Core Principle & Typical Service Workflow A professional service for Conventional SELEX typically follows this established, iterative cycle (8-15 rounds), as visualized below: Key Service Characteristics Target Immobilization: The target molecule is fixed to a solid support (e.g., magnetic beads, column resin, nitrocellulose membrane). Positive Selection: The library is passed over the immobilized target. Sequences with some binding affinity are retained, while others are washed away. Stringency Control: The service provider systematically increases selection pressure across rounds (e.g., by reducing target concentration, increasing wash stringency, adding counter-targets in Negative SELEX steps) to drive the evolution of high-affinity, specific binders. Monitoring: Enrichment is tracked via quantitative PCR, and final pools are analyzed by Next-Generation Sequencing (NGS) to identify convergent sequence families. Common Applications for this Service This classic approach is ideal for: Proteins that are stable and can be immobilized without losing native conformation (e.g., antibodies, enzymes, recombinant tags). Large molecules or complexes (e.g., viruses, whole cells—though Cell-SELEX is now more common for cells). Establishing proof-of-concept for a new target class where simpler, robust methodology is preferred. Deliverables Similar to other SELEX services, clients receive: A final report detailing the selection process and conditions. A list of top-ranked aptamer sequences with affinity (Kd) and specificity data. Aliquots of synthesized, validated…
What is Capture-SELEX? Unlike traditional SELEX where the target is immobilized, Capture-SELEX immobilizes the initial DNA library itself via a short complementary "capture" sequence. The key target molecule is free in solution. Binding occurs when an aptamer candidate in the library binds to the target, causing a structural change that releases it from the immobilization surface. This approach offers distinct advantages: Ideal for small molecules and proteins: Especially targets that are difficult to immobilize without affecting their structure. Minimizes non-specific binding: Selection pressure is purely for target-induced structure formation/release. Enriches for structure-switching aptamers: Resulting aptamers often undergo conformational change upon binding, making them excellent for biosensor development. Typical Capture-SELEX Screening Service Workflow A professional service provider will manage this complex, iterative process from start to finish. Here’s what you can expect: Phase 1: Project Design & Library Preparation Consultation & Target Specification: Defining target properties, desired affinity (Kd), specificity (against which counter-targets), and buffer conditions. Customized Library Design: Designing a single-stranded DNA library (10^14 - 10^15 unique sequences) with: A central random region (e.g., 30-50 nucleotides). Fixed primer regions for PCR amplification. A capture sequence region complementary to an immobilized oligonucleotide. Immobilization Matrix Preparation: Coupling the complementary "capture" oligonucleotides to a solid support (e.g., magnetic beads, chromatography resin). Phase 2: The Iterative Selection (SELEX) Cycles…
What is Filter Membrane Binding SELEX? SELEX (Systematic Evolution of Ligands by EXponential enrichment) is the standard method for discovering high-affinity, specific nucleic acid aptamers. The Filter Membrane Binding variant is one of the most classic and robust SELEX techniques. Core Principle: It leverages a nitrocellulose or mixed cellulose ester filter membrane, which irreversibly binds proteins and other macromolecules but allows short, unbound single-stranded DNA or RNA oligonucleotides to pass through. The Selection Mechanism: During each selection round, the target molecule (e.g., a protein) is immobilized on the filter. An immense library of random oligonucleotides (10^13 - 10^15 unique sequences) is applied. Only sequences that bind to the target are retained on the filter with it. Unbound sequences are washed away. The bound aptamer candidates are then eluted, amplified by PCR (or RT-PCR for RNA), and used as the enriched library for the next round. Key Features of the Service A professional service will typically offer: Target Flexibility: Optimal for purified proteins (recombinant or native), protein complexes, viruses, and even some small molecules if conjugated to a carrier protein. Counter-SELEX: A critical step to ensure specificity. The enriched library is passed through a filter bound to non-target molecules (e.g., related proteins, cell lysates, immobilization matrix) to subtract cross-reactive binders. High-Throughput…
What is SELEX and What are Aptamers? Aptamers: Often called "chemical antibodies," they are short, single-stranded DNA or RNA oligonucleotides that fold into specific 3D shapes to bind with high affinity and specificity to a target molecule (e.g., a viral protein, whole bacterium, or parasite surface marker). SELEX (Systematic Evolution of Ligands by EXponential enrichment): This is the iterative combinatorial chemistry process used to discover aptamers from a vast random library (10^14-10^15 unique sequences). It involves repeated cycles of: 1) Binding the library to the target, 2) Separating bound from unbound sequences, 3) Amplifying the bound sequences, and 4) Starting a new, enriched cycle. Core Components of a Pathogen SELEX Service A professional service will typically manage the entire pipeline: 1. Project Design & Target Preparation: Consultation: Defining the precise target (e.g., whole inactivated SARS-CoV-2, Salmonella outer membrane protein, Plasmodium lysate). Counter-SELEX: A critical step for pathogen specificity. The process is run against related non-targets (e.g., host cells, non-pathogenic bacterial strains) to filter out cross-reactive aptamers, ensuring the final aptamers distinguish between pathogen and non-pathogen. 2. The SELEX Execution: Performing multiple (usually 8-15) rounds of the selection process under optimized conditions (buffer, temperature, washing stringency). 3. Next-Generation Sequencing (NGS) & Bioinformatics: After the final rounds, the enriched pool is sequenced using NGS. Bioinformatic analysis identifies sequence…
What is a Stem Cell Aptamer Screening Service? It is a contract research service where a specialized lab uses Systematic Evolution of Ligands by EXponential Enrichment (SELEX) to discover and develop DNA or RNA aptamers that bind with high affinity and specificity to a target of your choice related to stem cells. Aptamers are often called "chemical antibodies." They are short, single-stranded oligonucleotides that fold into unique 3D shapes, allowing them to bind to targets like proteins, small molecules, or even whole cells. Core Targets for Stem Cell Applications The service can be tailored to screen for aptamers against: Specific Cell Surface Markers: (e.g., CD34, CD133, SSEA-4, TRA-1-60) for identification and isolation. Whole Live Stem Cells: To get aptamers that recognize the unique molecular signature of a specific stem cell type (e.g., mesenchymal stem cells, cancer stem cells, pluripotent stem cells). Differentiation State-Specific Targets: To distinguish between pluripotent, progenitor, and fully differentiated cells. Specific Stem Cell-Derived Products: (e.g., exosomes, vesicles). Typical Workflow of the Service A professional service provider will guide you through these stages: Phase Description Your Input 1. Project Design Defining the target (specific protein, cell line, primary cells), counter-selection cells (to ensure specificity), and desired aptamer properties (e.g., Kd, nuclease resistance). Provide target cells, control cells, and…
What is a Bacterial Aptamer Screening Service? It is a specialized contract research service where a provider uses Systematic Evolution of Ligands by Exponential Enrichment (SELEX) to discover and develop single-stranded DNA or RNA aptamers that bind with high affinity and specificity to a bacterial target. The target can be: Whole bacterial cells (e.g., E. coli O157:H7, Salmonella typhimurium). Specific bacterial components (e.g., surface proteins like pili, flagella, capsular polysaccharides, secreted toxins). Key virulence factors (e.g., endotoxins like LPS). The resulting aptamers are powerful recognition elements for diagnostics, therapeutics, and research. Core Steps in the Service Pipeline A typical full-service offering includes: 1. Project Design & Target Preparation: Consultation: Defining the goal (e.g., detection of a specific strain, therapeutic neutralization). Target Choice: Deciding between whole cells (for broad detection) or purified components (for precise targeting). Counter-SELEX: Using related non-target cells (e.g., non-pathogenic strain) to eliminate cross-reactive aptamers and ensure specificity. 2. Library Synthesis & SELEX Process: Library Design: Using a random-sequence oligonucleotide library (typically ~10^14 different molecules). Selection Rounds (8-15 cycles): Iteratively incubating the library with the target, washing away unbound sequences, eluting the bound ones, and amplifying them via PCR (for DNA) or RT-PCR (for RNA). Monitoring: Using quantitative PCR or flow cytometry to track enrichment progress. 3. Next-Generation Sequencing (NGS) & Bioinformatics:…
