PMID: 7519938May 1, 1994Paper

Large-scale preparation of highly purified, frozen/thawed CD34+, HLA-DR- hematopoietic progenitor cells by sequential immunoadsorption (CEPRATE SC) and fluorescence-activated cell sorting: implications for gene transduction and/or transplantation

Bone Marrow Transplantation
M KörblingM Andreeff

Abstract

The purification of early hematopoietic progenitor cells for autologous transplantation is based on two rationales: (1) elimination of clonogenic tumor cells, and/or (2) gene transfer into indefinitely self-replicating hematopoietic stem cells. Primitive CD34+ stem cells can be separated from more mature stem cells, or probably from clonogenic tumor cells, by differences in HLA-DR surface antigen expression. The objective of this study was to establish a large-scale technique for purification of CD34+, DR- progenitor cells from a large volume marrow harvest. In five different experiments, CD34+ cells were purified to between 76% and 91% by avidin-biotin immunoadsorption (CEPRATE SC) as a first step. This was followed by fluorescence-activated cell sorting to separate DR+ and DR- cells, which resulted in the generation of between 1.75 and 11.3 x 10(5) CD34+, DR- cells. The purity of DR- cells increased from between 0.5% and 4.3% in the immuno-adsorbed fraction up to 99% in the DR- sorted fraction. As shown in a single experiment, the purity of CD34+, DR- cells immediately after thawing increased from 0.01% to 94.3% while losing 99% of those early progenitor cells during the multistep purification procedure. We were able to physi...Continue Reading

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