Strategies for Isolating and Enriching Cancer Stem Cells: Well Begun Is Half Done

Abstract

Cancer stem cells (CSCs) constitute a subpopulation of cancer cells that have the potential for self-renewal, multipotent differentiation, and tumorigenicity. Studies on CSC biology and CSC-targeted therapies depend on CSC isolation and/or enrichment methodologies. Scientists have conducted extensive research in this field since John Dick's group successfully isolated CSCs based on the expression of the CD34 and CD38 surface markers. Progress in CSC research has been greatly facilitated by the enrichment and isolation of these cells. In this review, we summarize the current strategies used in our and other laboratories for CSC isolation and enrichment, including methods based on stem cell surface markers, intracellular enzyme activity, the concentration of reactive oxygen species, the mitochondrial membrane potential, promoter-driven fluorescent protein expression, autofluorescence, suspension/adherent culture, cell division, the identification of side population cells, resistance to cytotoxic compounds or hypoxia, invasiveness/adhesion, immunoselection, and physical property. Although many challenges remain to be overcome, it is reasonable to believe that more reliable, efficient, and convenient methods will be developed in the near future.

Characterization of HIF‐1α/Glycolysis Hyperactive Cell Population via Small‐Molecule‐Based Imaging of Mitochondrial Transporter Activity

“…Cancer stem‐like cells are defined as a subset of cancer cells that retain extensive self‐renewal potential and have the ability to recreate the heterogeneity of the original tumor 17. To confirm whether IR‐780 H cancer cells possess tumor initiating ability, human lung cancer cells A549 were labeled with IR‐780 and sorted using flow cytometry as previous described.…”

Progress in isolation and enrichment of cancer stem cells

“…The characteristics of this model is adding EGF, bFGF growth factor in the medium. This uncomplicated and convenient method for NSCs' enrichment has been extensively applied to study neurogenesis and also been applied for CSCs' isolation from brain tumors and other types of cancers with different combinations of complementary agents [16] . Many scholars applied the method of serum-free culture to isolate breast cancer stem cells from breast cancer [17][18] .…”

Therapeutic strategies for targeting cancer stem cells

“…This new marker has been proven to be a more reliable and accurate way to identify and characterize CSCs. [16,23] Another important functional property of CSCs, as well as normal stem SCs, is the ability to produce sphere-forming colonies from a single cell in serum-free medium or in soft agar medium, as differentiated cells cannot survive and proliferate in this environment. [24] Thus, several studies have used the sphere formation assay as an efficient method for isolating, enriching and maintaining CSCs from various primary tumors.…”

“…[18,25] These spheres clearly demonstrated stem-like properties and expressed characteristics of CSCs. [16] Here, we will focus on cancer therapeutics which can target CSCs. The development of various strategies that can act effectively against CSCs has been categorized into six groups, as shown in Figure 1.…”

“…They can be identified, isolated, and characterized by several methodologies, including flow cytometric analysis of CSC-specific cell surface markers, detection of side-population (SP) phenotypes by Hoechst 33342 dye exclusion, changes in aldehyde dehydrogenase (ALDH) enzymatic activities using an aldeflour assay, ability to grow as suspension spheres in serum-free medium, SC-related gene expression, and auto-fluorescence. [6,[15][16][17] There are no widely accepted techniques solely developed to isolate CSCs, necessitating the use of combination markers and methods rather than single strategies.…”