Protocol for sorting cancer stem cells using a combination of anti-CD133 antibody and lectin cyanovirin-N

• Published in: STAR protocols 2024-12, Vol.5 (4), p.103443, Article 103443
• Main Authors: Wei, Yuanyan, Geng, Shuting, Chen, Qihang, Chen, Jiayue, Fan, Zhijun, Li, Yi, Si, Yu, Yang, Yuerong, Liang, Ziwei, Jiang, Jianhai
• Format: Article
• Language: English
• Subjects: AC133 Antigen - metabolism; Antibodies - immunology; Bacterial Proteins - metabolism; cancer; Carrier Proteins - metabolism; cell biology; cell isolation; Cell Line, Tumor; Cell Separation - methods; Flow Cytometry - methods; Humans; Neoplastic Stem Cells - metabolism; Pore Forming Cytotoxic Proteins - metabolism; Protocol; stem cells
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2024.103443

CD133 is widely used as a marker to isolate cancer stem cells (CSCs). However, the structural ambiguity of N-glycan of CD133 limits its application in the isolation of CSCs. Here, we present a protocol to sort CSCs from tumor samples by combining CD133 with α-1,2-high-mannose type glycan chains. We describe steps for purifying and biotinylating cyanovirin-N (CVN) proteins and sorting tumor cells and CD133+α-1,2-Man+ (α-1,2-linked mannose+) cells to provide more specific markers for CSCs. For complete details on the use and execution of this protocol, please refer to Wei et al.1 [Display omitted] •Detailed protocol for culturing primary cells of intrahepatic cholangiocarcinoma•Sorting cancer stem cells from intrahepatic cholangiocarcinoma tumor samples•α-1,2-Mannosylated CD133 is a marker for sorting and identifying cancer stem cells Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. CD133 is widely used as a marker to isolate cancer stem cells (CSCs). However, the structural ambiguity of N-glycan of CD133 limits its application in the isolation of CSCs. Here, we present a protocol to sort CSCs from tumor samples by combining CD133 with α-1,2-high-mannose type glycan chains. We describe steps for purifying and biotinylating cyanovirin-N (CVN) proteins and sorting tumor cells and CD133+α-1,2-Man+ (α-1,2-linked mannose+) cells to provide more specific markers for CSCs.