Human hematopoietic cell culture, transduction, and analyses.
This unit provides methods for introducing genes into human hematopoietic progenitor cells. The Basic Protocol describes isolation of CD34(+) cells, transduction of these cells with a retroviral vector on fibronectin-coated plates, assaying the efficiency of transduction, and establishing long-term cultures. Support protocols describe methods for maintenance of vector-producing fibroblasts (VPF) and supernatant collection from these cells, screening medium components for the ability to support hematopoietic cell growth, and establishing colonies from long-term cultures. Other protocols provide PCR-based methods to analyze individual colonies for transduction, methods to analyze cells harvested from long-term cultures, and procedures for freezing and thawing of hematopoietic cells.
Bonde
Jesper
J
Statens Serum Institut, Copenhagen, Denmark.
Wirthlin
Louisa
L
Kohn
Donald B
DB
Nolta
Jan A
JA
D018952:Antigens, CD34; D018929:Cell Culture Techniques; D003584:Cytological Techniques; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D014161:Transduction, Genetic
D018952:Antigens, CD34