Preparation of Immunomagnetic Microspheres and Their Application in Selection of Murine Hematopoietic Stem Cells

• Main Authors: Jing-yi Chang, 張靜宜
• Other Authors: Wen-chien Lee
• Format: Others
• Language: zh-TW
• Published: 2007
• Online Access: http://ndltd.ncl.edu.tw/handle/01369025261191590416

碩士 === 國立中正大學 === 化學工程所 === 95 === Hematopoietic stem cells (HSCs) are widely used in leukemia，lymphoma，inherited blood disorders，gene therapy and so on . Very rare amount of HSCs is enough to save life of lethally irradiated patients. However the amounts of HSCs are very rare in the body. About one in every 10,000 to 15,000 bone marrow cells is thought to be a HSC. In the blood stream the proportion falls to one in 100,000 blood cells.The isolation and purification of HSCs is thus of great importance in both research and clinical applications. Both indirect and direct immunomagnetic methods for separating HSCs in the suspension of MNCs from mice（BALB/Cs）femurs and tibias were carried out based on the positive selection of Sca-1 surface marker (Sca-1+). In the indirect method，via use of purified anti-mouse Sca-1 antibody were bound onto magnetic micro beads coupled with 15 mg/g goat anti rat antibody to form immunomagnetically labeled cell .The optimal protocol of indirect immunomagnetic separation consisted of a bead-to-cell ratio of 10:1 and a mice age of 6 weeks. The highest purities of Sca-1+ and c-Kit+ were 89.4 and 92.9 and the enrich fold of Sca-1+ was 19.6 when the beads-to-cell ratio was at 10:1. In the direct method，after the coupling of biotin anti-mouse Sca-1 antibody bound magnetic microbeads could catch target cells (Sca-1+) to form immunomagnetically labeled cell .The highest purities of Sca-1+ and c-Kit+ were 85.0 and 85.4 and the enrich fold of Sca-1+ was 16.3 when the bead-to-cell ratio was at 10:1.