“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…
Core Concept & Purpose The goal is to subtract sequences that bind to: The immobilization matrix/surface (e.g., streptavidin beads, nitrocellulose filters, chip surface). Closely related molecules or structural analogs (e.g., to ensure an aptamer for drug A doesn't bind metabolite B). Components of the selection buffer or the cellular milieu where the aptamer will be used (e.g., serum proteins for therapeutic aptamers). By pre-incubating the DNA library with these "negative targets" before exposure to the desired target, non-specific binders are captured and discarded. Only the unbound, "cleaned" library proceeds to the positive selection round. Key Features of a Professional Negative SELEX Service Strategic Design: Experts design the optimal order, frequency, and stringency of negative vs. positive selection rounds. Relevant Negative Targets: The service advises on and sources the most critical counter-targets (e.g., using the exact resin from positive selection for matrix subtraction, or sourcing specific protein analogs). Dedicated Rounds: Entire selection rounds may be dedicated to negative selection against a key interferent. Pre-SELEX Depletion: Often, the initial naive library is pre-depleted against the matrix to remove common surface binders from the start. Typical Integration into a Service Workflow Within a broader SELEX project (e.g., Protein-SELEX), a Negative SELEX step is woven in as follows: Pre-Clearance (Round 0): The initial DNA library is…
Core Concept: SELEX SELEX (Systematic Evolution of Ligands by Exponential Enrichment) is an iterative process to select high-affinity, specific nucleic acid aptamers (ssDNA or RNA) against a target molecule. The magnetic bead-based method revolutionizes this by using beads as a solid, easily separable support, drastically improving speed and efficiency. How Magnetic Bead-Based SELEX Works (The Process) A professional service provider will execute this cyclic process, usually over 8-15 rounds: Key Advantages of the Magnetic Bead Method (Why it's the service of choice) Rapid Separation: Magnetic stands enable quick washing and buffer exchange, shortening each selection round from hours to minutes. Reduced Non-Specific Binding: Efficient washing minimizes background, leading to cleaner selection. Automation-Friendly: Perfectly suited for robotic liquid handlers, enabling high-throughput, reproducible selections. Flexibility with Targets: Immobilized Targets: Proteins, small molecules, cells, or viruses can be directly conjugated to the beads. Counter-Selection: Beads coated with non-target molecules (e.g., a related protein, cell type) can be used to subtract non-specific binders, dramatically enhancing specificity. What a Full-Service Provider Typically Offers 1. Project Design & Consultation Target characterization and strategy (native vs. tagged protein, cell surface marker). Selection of library type (DNA, RNA, modified nucleotides like 2'-F for stability). Design of counter-selection steps. 2. The SELEX Selection Process Library Synthesis: Starting with…
