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EC-Cult™-XF ECFC Culture Kit

Culture kit for derivation and proliferation of human endothelial colony-forming cells

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EC-Cult™-XF ECFC Culture Kit

Culture kit for derivation and proliferation of human endothelial colony-forming cells

1 Kit
Catalog #08000
599 USD

Required Products

Overview

EC-Cult™-XF ECFC Culture Kit includes EC-Cult™-XF ECFC Basal Medium, EC-Cult™-XF ECFC 2.5X Supplement, and Animal Component-Free Cell Attachment Substrate. It is formulated to derive, maintain, and differentiate human endothelial colony-forming cells (ECFCs). Complete EC-Cult™-XF ECFC Medium is developed to derive primary human ECFCs from human umbilical cord blood and human adult peripheral blood, support long-term expansion, and maintain clonogenic properties along the entire ECFC hierarchy. This kit is optimized for isolating and counting ECFCs using a 7- to 14-day ECFC assay.

Complete EC-Cult™-XF ECFC Medium must be supplemented with Heparin Solution (Catalog #07980). For passaging ECFCs, Animal Component-Free Cell Dissociation Kit (Catalog #05426) is required.
Advantages:
• Supports derivation, expansion and maintenance of ECFCs in a xeno-free culture system
• Supports greater long-term expansion of ECFCs than commercially available or serum-containing media and maintains endothelial cell surface markers and morphology
• Supports clonogenic expansion of ECFCs while maintaining proliferative potential for greater than 10 passages
•Rigorous raw material screening and quality control minimize lot-to-lot variability and increase reproducibility between experiments
•ECFCs cultured in Complete EC-Cult™-XF ECFC Medium demonstrate ex vivo and in vivo functionality
Components:
• EC-Cult™-XF ECFC Basal Medium, 300 mL
• EC-Cult™-XF ECFC 2.5X Supplement, 200 mL
• Animal Component-Free Cell Attachment Substrate, 1 mL
Subtype:
Specialized Media
Cell Type:
Angiogenic Cells; Endothelial Cells
Species:
Human
Application:
Cell Culture
Brand:
EC-Cult
Area of Interest:
Angiogenic Cell Research; Endothelial Cell Biology; Stem Cell Biology
Formulation:
Xeno-Free

Scientific Resources

Educational Materials

(2)

Data and Publications

Data

Derivation and Colony Assay for Human ECFCs Using EC-Cult™-XF ECFC Medium

Figure 1. Derivation and Colony Assay for Human ECFCs Using EC-Cult™-XF ECFC Medium

(A) A schematic of the ECFC assay for enumerating ECFC colonies derived in Complete EC-Cult™-XF ECFC Medium. On Day 0, mononuclear cells are isolated from whole blood and plated on animal component-free (ACF) Cell Attachment Substrate-coated plates in Complete EC-Cult™-XF ECFC Medium. From Day 1 to 7, nonadherent cells are gently removed through daily medium changes. From Day 8 and onwards, the medium is changed every second day until ECFCs are ready to be passaged. Each ECFC colony can be enumerated around Day 10 before the colonies begin to merge onto each other. (B) A representative phase contrast image of an ECFC colony at Day 11 derived from human umbilical cord blood (CB) in Complete EC-Cult™-XF ECFC Medium. (C) A representative phase contrast image at passage 3 of ECFCs derived and expanded in Complete EC-Cult™-XF ECFC Medium. These cells grow as monolayers and retain their typical cobblestone-like morphology of endothelial cells.

Greater Expansion of Human ECFCs is Obtained When Using the EC-Cult™-XF ECFC Culture Kit

Figure 2. Greater Expansion of Human ECFCs is Obtained When Using the EC-Cult™-XF ECFC Culture Kit

Human CB-derived ECFCs expanded in Complete EC-Cult™-XF ECFC Medium demonstrate greater expansion rate when compared to ECFCs expanded in a Commercial Medium. In late passages, cells cultured in Complete EC-Cult™-XF ECFC Medium continue to expand at the same rate as in early passages, while the rate of expansion in cultures expanded in a Commercial Medium begins to decline as early as passage 6 (n = 4).

Clonogenic Potential of ECFCs Derived and Expanded in Complete EC-Cult™-XF ECFC Medium is Maintained

Figure 3. Clonogenic Potential of ECFCs Derived and Expanded in Complete EC-Cult™-XF ECFC Medium is Maintained

Human CB-derived ECFCs expanded and plated starting with one cell per well in EC-Cult™-XF ECFC Medium or Commercial Medium form colonies of varying sizes over a 14 day period. Assessment of colony formation and size is used to determine their hierarchical proliferative potential. The assessment includes counting and classifying the number of cells per colony as endothelial clusters (2-50 cells per colony), low proliferative (51-500 cells per colony), medium proliferative (501-2000 cells per colony) and high proliferative (greater than 2000 cells per colony) ECFCs. In Complete EC-Cult™-XF ECFC Medium, approximately 40 to 60% of dividing ECFCs form well-circumscribed colonies containing greater than 2000 cells at the end of the 14-day culture. A greater proportion of high proliferative ECFCs are maintained when ECFCs are derived and expanded in Complete EC-Cult™-XF ECFC Medium. Approximately 2% of the colonies are high proliferative when cultured in a commercial medium. Vertical lines indicate standard error (n=3).

Human ECFCs Culture-expanded in EC-Cult™-XF ECFC Medium are Homogeneous and Maintain High Expression of Endothelial Cell Surface Markers

Figure 4. Human ECFCs Culture-expanded in EC-Cult™-XF ECFC Medium are Homogeneous and Maintain High Expression of Endothelial Cell Surface Markers

ECFCs derived and expanded for 5 passages in EC-Cult™-XF ECFC Medium (A) maintain the expression of classic ECFC and endothelial cell surface markers (CD31 and CD144) and (B) lack the expression of hematopoietic marker, CD45.

ECFCs derived in EC-Cult™-XF ECFC Medium Form Tube-like Structures in Vitro

Figure 5. ECFCs derived in EC-Cult™-XF ECFC Medium Form Tube-like Structures in Vitro

ECFCs derived and expanded for 5 passages in EC-CultTM-XF ECFC Medium are able to spontaneously assemble into capillary tube-like structures overnight on Matrigel®.

Publications

(2)
Stem cells translational medicine 2017 MAY

Endothelial Progenitors: A Consensus Statement on Nomenclature.

Medina RJ et al.

Abstract

Endothelial progenitor cell (EPC) nomenclature remains ambiguous and there is a general lack of concordance in the stem cell field with many distinct cell subtypes continually grouped under the term EPC." It would be highly advantageous to agree on standards to confirm an endothelial progenitor phenotype and this should include detailed immunophenotyping potency assays and clear separation from hematopoietic angiogenic cells which are not endothelial progenitors. In this review we seek to discourage the indiscriminate use of "EPCs and instead propose precise terminology based on defining cellular phenotype and function. Endothelial colony forming cells and myeloid angiogenic cells are examples of two distinct and well-defined cell types that have been considered EPCs because they both promote vascular repair, albeit by completely different mechanisms of action. It is acknowledged that scientific nomenclature should be a dynamic process driven by technological and conceptual advances; ergo the ongoing EPC" nomenclature ought not to be permanent and should become more precise in the light of strong scientific evidence. This is especially important as these cells become recognized for their role in vascular repair in health and disease and in some cases progress toward use in cell therapy. Stem Cells Translational Medicine 2017;6:1316-1320.
JCI insight 2017 JAN

CD44 expression in endothelial colony-forming cells regulates neurovascular trophic effect.

S. Sakimoto et al.

Abstract

Vascular abnormalities are a common component of eye diseases that often lead to vision loss. Vaso-obliteration is associated with inherited retinal degenerations, since photoreceptor atrophy lowers local metabolic demands and vascular support to those regions is no longer required. Given the degree of neurovascular crosstalk in the retina, it may be possible to use one cell type to rescue another cell type in the face of severe stress, such as hypoxia or genetically encoded cell-specific degenerations. Here, we show that intravitreally injected human endothelial colony-forming cells (ECFCs) that can be isolated and differentiated from cord blood in xeno-free media collect in the vitreous cavity and rescue vaso-obliteration and neurodegeneration in animal models of retinal disease. Furthermore, we determined that a subset of the ECFCs was more effective at anatomically and functionally preventing retinopathy; these cells expressed high levels of CD44, the hyaluronic acid receptor, and IGFBPs (insulin-like growth factor-binding proteins). Injection of cultured media from ECFCs or only recombinant human IGFBPs also rescued the ischemia phenotype. These results help us to understand the mechanism of ECFC-based therapies for ischemic insults and retinal neurodegenerative diseases.
STEMCELL TECHNOLOGIES INC.’S QUALITY MANAGEMENT SYSTEM IS CERTIFIED TO ISO 13485. PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED.