What is Cell-SELEX?
Cell-SELEX is an exciting technique for the creation of aptamer affinity agents without prior identification of a specific target. (Learn about aptamers and traditional SELEX) Aptamers are selected based on surface differences between normal and cancer or disease cells and between different types of cancer cells (2,3). Aptamers can also be selected for cell internalization (4). Aptamers to multiple cell surface targets can be discovered in a single cell-SELEX process (1,3).
Traditional Cell-SELEX Process
Step 1: Bind Library to Negative Cells and Collect Supernatant Containing Non-Binders
Step 2: Bind Library to Positive Cells and Discard Supernatant Containing Non-Binders
Step 3: Isolate Positive Binders and Amplify via PCR
Step 4: Repeat Steps 1 through 3 using Amplified Positive Binders
As with standard aptamer SELEX, multiple rounds of selection (steps 1 through 3) are conducted. Enrichment of aptamer sequences is evaluated between rounds to determine when SELEX is complete.
Advantages of Cell-SELEX
Cell-SELEX offers several unique advantages. To start, aptamers can be selected without prior knowledge of a specific target. This can dramatically reduce both the time and cost to develop selective affinity reagents. In cell-SELEX, targets are presented in their natural conformation with natural modifications at their natural frequency on the cell surface. This can yield aptamers with enhanced affinity and selectivity for cell-surface targets. While aptamers to specific cells can be directly used in diagnostics, therapeutics and cell targeting, they can also be used to capture and identify novel aptamer targets. As early as 2008, researchers described a process for selecting aptamers to specific cells, then isolating and characterizing the aptamer targets (1).
Discovering Biomarkers with Aptamers
Disease diagnosis and drug development rely on specific targets, or biomarkers, that discriminate between normal and diseased cells. Identifying early indicators of disease is a time-consuming process. (4). For diseases lacking diagnostic biomarkers and therapeutic targets, aptamers are providing new hope. Unlike antibodies, which are typically raised against a purified target protein, aptamers can be selected for binding to whole cells. By selecting for aptamers that bind to a specific type of cancer cell, but do not bind to normal cells, researchers are discovering aptamers that exclusively target diseased cells. While these aptamers can sometimes be used in diagnostics and as direct therapeutic or drug delivery/cell targeting agents, aptamer binding can be analyzed to reveal novel biomarkers. Biomarkers discovered through cell-SELEX can be used for production of more traditional diagnostic and therapeutic agents.
Isolating and Characterizing Unknown Biomarkers following Cell-SELEX
Step 1: Bind Selective Aptamers to Target Cells
Step 2: Isolate Aptamer-labeled Cells using a Magnet
Step 3: Lyse Cells and Isolate Aptamer-Biomarker Complexes
Step 4: Denature Complexes and Collect Cell-Specific Biomarkers
Step 5: Trypsinize and Perform LC-MS to Identify Biomarkers
Cancer Biomarker Discovery Using Cell-SELEX
Researchers at Lanzhou University in China used cell-CELEX to identify a biomarker for early stage gastric cancer. A mixed cell population from individual donors with early-stage gastric cancer was used for positive aptamer selection. Normal gastric mucosa was used for negative selection. After 12 rounds of cell-SELEX, aptamer Ap7 was selected based on affinity and selectivity. Binding of fluorescein-labeled aptamer was used to confirm selectivity for early stage gastric cancer cells. Further analysis was performed to characterize the aptamer target. Based on MALDI-TOF mass spec and the NCBI database, the target protein was identified as peroxiredoxin-4. ELISA analysis confirmed higher expression of peroxiredoxin-4 in cells from early-stage gastric cancer than late-stage gastric cancer and normal gastric mucosa (6).
Custom Aptamer Discovery Using Cell-SELEX
Cell-SELEX is an exciting technique that enables selection of aptamers without prior identification of a specific cell surface target. Selection of aptamer affinity reagents to cells (and unknown targets) of interest is used to identify new disease-specific targets for therapeutic development and early-stage diagnostics. Combining traditional SELEX with cell-SELEX helps ensure selectivity for cell surface targets in their native form. Base Pair has performed a number of cell-SELEX projects, including selection of aptamers to cell-surface markers and identification of aptamers that enter specific cells.
Contact Base Pair today to learn more about aptamer selection to the specific cell line or cell population of interest to you.
- Berezovski, M.V., et al. Aptamer-facilitated biomarker discovery (Apta BiD). JACS Articles. 2008. 130:9137-9143.
- Kaur, H., et al. Recent developments in cell-SELEX technology for aptamer selection. Biochimica et Biophysica Acta. 2018. 1862(10):2323-2329.
- Li, W.M., et al. Selection of metastatic breast cancer cell-specific aptamers for the capture of CTCs with a metastatic phenotype by cell-SELEX. Molecular Therapy: Nucleic Acids. 2018. 12:707-717.
- Pang, X., et al. Bioapplication of cell-SELEX-generated aptamers in cancer diagnostics, therapeutics, theranostics, and biomarker discovery: a comprehensive review. Cancers. 2018. 10(2):47
- Wu, Q. DNA aptamer from whole cell-SELEX as new diagnostic agents against glioblastoma multiforme cells. 2018. 143:2267-2275.
- Xu, P., et al. Peroxiredoxin-4 as a potential biomarker of early gastric cancer screened by cell-SELEX. Translational Cancer Research. 2017. 6(2)