Derivation of primary cell cultures from solid tumor material involves multiple critical handling steps. This workflow improves this complex procedure. It allows for rapid and flexible tissue preparation and for subsequent derivation and expansion of tumor cell cultures from primary tumors.
The workflow preserves the parental tumor heterogeneity, tumor-initiating capacity and genetic stability of the cell culture. This leads to improved in vitro models for cancer research and drug screening. Moreover, stable cell lines from primary human tumor tissue can be generated.
Derivation of primary cell cultures from solid tumor material involves multiple critical handling steps. To improve this complex procedure, we developed a workflow allowing for rapid and flexible tissue preparation for subsequent derivation of primary tumor cell cultures using our novel TumorMACS™ Media.
The workflow enables highly standardized cell line derivation and still offers the opportunity to seed dissociated tumors and/or selected cell fractions such as isolated tumor cells or tumor subpopulations.
The gentleMACS™ Dissociator in combination with the Tumor Dissociation Kit, human ensures rapid and standardized dissociation of tissue samples into single-cell suspensions. No matter the tumor entity, gentleMACS Technology provides excellent cell viability and functionality for further downstream cell separation, analysis, and culture.
The presence of red blood cells (RBCs) hampers the accurate determination of cell viability as well as absolute cell counts for tumor-infiltrating leukocytes, cancer stem cells, and other tumor-resident cells. Labeling the cells with CD235a (glycophorin A) antibody allows for the fast and simple discrimination and exclusion of RBCs for subsequent flow cytometric analysis.
The Pancreas, Renal, and Ovarian TumorMACS™ Media have been developed for derivation and expansion of human tumor cell cultures from respective tumor entities. The media ares optimized to generate primary cell cultures and cell lines from both primary patient tumors as well as xenotransplanted tumors. Properties of the generated cell cultures closely resemble the parental tumor, including expression of subtype-specific markers, cellular heterogeneity, genetic as well as epigenetic signatures, and tumorigenic potential.
Using TumorMACS™ Media, cells can be plated directly after dissociation or after tumor cell isolation and both, adherent and suspension cells can be successfully expanded. Primary cell lines resemble the parent tumor, as morphological heterogeneity is preserved even in late passages.
While cell line–derived xenografts show a homogeneous histology, primary cell lines derived from primary tumors and propagated in TumorMACS™ Media retain their initial heterogeneity. After xenotransplantation, these cell lines closely resemble the parental patient tumor in histological analyses and thus are valuable in vivo tumor models.
Long-term propagation of tumor cell lines in vitro harbors the risk of inducing bias in gene expression profiles, thus resulting in the divergence from parental tumors. In primary human cell lines propagated in TumorMACS™ Media, the global gene expression profile was maintained upon long-term cultivation as well as for primary cell line–derived xenotransplanted tumors.
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