What is CE-SELEX? SELEX (Systematic Evolution of Ligands by EXponential Enrichment) is the standard process for aptamer development. It involves iterative rounds of selection and amplification to enrich nucleic acid sequences that bind tightly to a target molecule. Traditional SELEX often uses immobilization of the target on beads or filters, which can be slow (8-15 rounds) and may introduce bias by selecting for sequences that bind to the immobilization matrix itself. CE-SELEX uses Capillary Electrophoresis as the separation mechanism. The key principle is that when an aptamer binds to its target, it forms a complex with a different charge-to-size ratio, causing it to migrate at a different time (shifted peak) in the capillary compared to the unbound nucleic acid library. This complex can be isolated and collected with exquisite precision. Core Advantages of a CE-SELEX Screening Service A service provider offering CE-SELEX delivers significant benefits: Extreme Speed and Efficiency: Often requires only 2-4 rounds of selection to obtain high-affinity aptamers (nanomolar to picomolar Kd), compared to many more rounds in traditional SELEX. This translates to weeks or months of time saved. Solution-Phase Selection: The target is free in solution, eliminating immobilization bias. This allows for selection against targets in their native conformation and enables selection for small molecules and…
What is Whole-cell SELEX? Whole-cell SELEX (Systematic Evolution of Ligands by EXponential enrichment) is a technique used to discover aptamers (single-stranded DNA or RNA molecules) that bind specifically to a target living cell. Unlike traditional SELEX that uses a purified protein target, whole-cell SELEX presents the target in its native, complex cellular environment. This allows for the selection of aptamers against: Native cell-surface proteins in their proper folding and post-translational modifications. Complex targets like transmembrane receptors in their natural lipid environment. Unknown surface biomarkers without prior knowledge of the cell's molecular makeup. Specific cell states (e.g., activated, cancerous, infected) based on differences in surface expression. The Core Process: How Whole-cell SELEX Works A professional service will manage this complex, iterative pipeline: Library & Design: Starting with a vast, random synthetic oligonucleotide library (10^14 - 10^15 unique sequences). Positive Selection: Incubating the library with the target cells (e.g., cancer cells, stem cells, bacteria). Aptamers that bind to any surface structure are retained. Counter-Selection (Critical Step): The bound pool is then exposed to non-target or control cells (e.g., healthy cells, a different cell line). Sequences that bind to these are discarded. This step is crucial for generating specificity. Elution & Amplification: Aptamers specifically bound to the target cells are recovered, amplified by PCR…
Core Components of a Small Molecule Target Service A comprehensive service follows the early drug discovery workflow: 1. Target Identification & Prioritization Bioinformatics & Omics Analysis: Mining genomic, proteomic, and clinical data to find proteins or pathways dysregulated in a disease. Genetic Screens: Using CRISPR-Cas9 or RNAi to knock out/knock down genes and identify which are essential for disease cell survival. Literature & Database Mining: Systematic review of existing scientific and patent data to propose novel or repurposable targets. 2. Target Validation In Vitro Models: Confirming the target's role in disease using engineered cell lines (overexpression, knockout) and relevant disease models (e.g., cancer cell lines, neuronal cultures). In Vivo Models: Using animal models (e.g., zebrafish, mice) to see if modulating the target (genetically or with a tool compound) has the desired therapeutic effect and is safe. Biochemical Validation: Demonstrating the target protein is expressed, has the expected activity, and is "druggable" (has a pocket where a small molecule can bind). 3. Assay Development & Screening This is a critical service. Providers develop robust tests ("assays") to measure target activity. Types of Assays: Biochemical Assays: Test compound binding/interaction with the purified target protein (e.g., enzymatic activity, protein-protein interaction). Cell-Based Assays: Test compound function in a living cell (e.g., reporter…
Core Concept The central idea is "Target-based Drug Discovery." Instead of screening compounds on whole cells or organisms (phenotypic screening), you start with a specific protein (e.g., a kinase, receptor, ion channel) implicated in a disease. Services then help you understand that target and find molecules that modulate it. Categories of Protein Target Services These services typically follow the drug discovery pipeline: 1. Target Identification & Validation Bioinformatics & Omics Analysis: Mining genomic, proteomic, and clinical data to identify novel disease-associated targets. Genetic Validation: CRISPR/Cas9 gene editing (knock-out/knock-in), siRNA/shRNA knockdown to confirm the target's role in disease pathways. Functional Validation: Cell-based assays to see if modulating the target affects disease-relevant phenotypes. 2. Protein Expression & Purification Recombinant Protein Production: Cloning, expressing (in E. coli, insect, or mammalian cells), and purifying milligram to gram quantities of the target protein. This is essential for structural studies and biochemical assays. Membrane Protein Expertise: Specialized services for difficult-to-express targets like GPCRs and ion channels. Tagging & Labeling: Adding tags (His, GST, FLAG) for purification or fluorescent/isotopic labels for assays. 3. Structural Biology & Biophysics X-ray Crystallography: Determining high-resolution 3D structures of protein-ligand complexes. Cryo-Electron Microscopy (Cryo-EM): For large complexes or membrane proteins unsuitable for crystallography. Nuclear Magnetic Resonance (NMR) Spectroscopy: For studying dynamics and ligand binding in solution. Surface…
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…
Aptamers are single-stranded oligonucleotides that fold into defined architectures and bind to targets such as proteins. In binding proteins they often inhibit protein–protein interactions and thereby may elicit therapeutic effects such as antagonism. Aptamers are discovered using SELEX (systematic evolution of ligands by exponential enrichment), a directed in vitro evolution technique in which large libraries of degenerate oligonucleotides are iteratively and alternately partitioned for target binding. They are then amplified enzymatically until functional sequences are identified by the sequencing of cloned individuals. For most therapeutic purposes, aptamers are truncated to reduce synthesis costs, modified at the sugars and capped at their termini to increase nuclease resistance, and conjugated to polyethylene glycol or another entity to reduce renal filtration rates. The first aptamer approved for a therapeutic application was pegaptanib sodium (Macugen; Pfizer/Eyetech), which was approved in 2004 by the US Food and Drug Administration for macular degeneration. Eight other aptamers are currently undergoing clinical evaluation for various haematology, oncology, ocular and inflammatory indications. Aptamers are ultimately chemically synthesized in a readily scalable process in which specific conjugation points are introduced with defined stereochemistry. Unlike some protein therapeutics, aptamers do not elicit antibodies, and because aptamers generally contain sugars modified at their 2′-positions,…
What is an Aptamer? An aptamer is a short, single-stranded oligonucleotide (DNA or RNA) or peptide that binds to a specific target molecule (e.g., proteins, small molecules, cells, viruses) with high affinity and specificity. Often called "chemical antibodies," they offer advantages like stability, low-cost synthesis, and minimal batch-to-batch variation. The Core Process: SELEX The standard method for aptamer selection is SELEX (Systematic Evolution of Ligands by EXponential enrichment). Basic SELEX Workflow: Library Synthesis: Create a vast random-sequence oligonucleotide library (typically 10¹³ - 10¹⁵ unique sequences) flanked by constant primer regions for PCR amplification. Incubation: The library is incubated with the target molecule under controlled conditions (buffer, temperature, time). Partitioning: Bound sequences are separated from unbound ones. This is the most critical step and varies based on target (e.g., filtration, affinity columns, magnetic bead separation). Elution: Bound aptamers are recovered from the target (e.g., by denaturation or competitive elution). Amplification: The recovered pool is amplified by PCR (for DNA) or RT-PCR (for RNA) to create an enriched library for the next round. Iteration: Steps 2-5 are repeated (typically 8-15 rounds) to progressively enrich for sequences with the highest affinity and specificity. Cloning & Sequencing: The final enriched pool is cloned and sequenced to identify individual aptamer candidates. Key Variants of…
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."…
