Embryonic Stem Cell FBS Qualified
Gibco Embryonic Stem (ES) Cell FBS Qualified uses an industry-leading qualification assay, which includes state-of-the-art instrumentation, cell culture, and assay design. The improved assay was developed by our Stem Cell R&D and Quality teams to confirm the ability of our FBS to achieve high plating efficiencies and maintain pluripotency of ES cells. The new method evaluates the ability of FBS to support plating, pluripotency and proliferation in mESCs by quantifying pluripotency marker expressing colonies in combination with morphological evaluation.
Advantages of using Gibco Embryonic Stem Cell FBS Qualified
- Specially tested for the ability to sustain undifferentiated ES cells (Figure 1A) while maintaining karyotype integrity (Figure 2), LIF responsiveness (Figure 3) and pluripotency markers (Figure 1B)
- New improved screening with germ line–competent PRX129/X1 mESC line using a predictive assay that measures plating efficiency and pluripotency maintenance
- High consistency between lots with proven applications in iPSC generation and PSC culture
- Now available in the aliquot-free One Shot 50 mL bottle designed specifically for FBS use
mESCs grown in ES Cell FBS Qualified exhibit pluripotency traits
mESCs grown in media supplemented with ES Cell FBS Qualified exhibit morphological characteristics of pluripotency: well-defined refractive edges, tight colonies with small cells, and few differentiated cells (Figure 1A). ESC colonies grown in ES Cell FBS Qualified also stain positive for alkaline phosphatase (AP). AP is a pluripotency marker expressed by PSCs.
Figure 1. Image analysis of mESCs colonies grown in media containing Gibco ES Cell FBS Qualified. (A) Phase-contrast image of mESC colonies grown in media containing ES Cell FBS Qualified showing colonies with morphological traits consistent with pluripotency. (B) An overlay of the fluorescence image (obtained after staining the sample with Alkaline Phosphatase (AP) Live Stain) with the phase-contrast image of mESC colonies, showing uniform AP staining. Colonies that appear green are expressing AP and are therefore pluripotent.
Figure 2. mESCs grown in media supplemented with ES Cell FBS Qualified exhibit normal karyotypes. Normal karyotype report for PRX129/X1 mESC line. 18 of 20 cells tested exhibited a normal karyotype. (Cell Line Genetics).
Figure 3. mESCs grown in media supplemented with ES Cell FBS Qualified maintain LIF responsiveness. When LIF is removed from mESC media, normal mESC cultures respond with widespread differentiation. PRX129/X1 murine ES cells grown with and without LIF demonstrate this response.
ES Cell FBS Qualified is able to support reprogramming workflows
Figure 4. When used to supplement growth media, ES Cell FBS Qualified is able to support generation of iPSCs from both mouse and human fibroblasts. In a standard workflow using Sendai virus–based reprogramming (CytoTune iPS 2.0 Sendai Reprogramming Kit), both murine and human fibroblasts successfully generated iPSCs.
Rigorous screening methodology helps ensure superior FBS performance
Each lot of ES Cell FBS Qualified is screened using PRX129/X1 murine ESC line using an assay that measures plating efficiency, morphology (Figure 5), and stem cell marker expression (Figure 6).
Figure 5. Plating efficiency and colony morphology assessed in murine ESCs grown in media containing ES Cell Qualified FBS. (A) Undifferentiated colonies (top row) appear bright and sharp, and differentiated colonies (bottom row) appear dim, fuzzy and flattened when stained with alkaline phosphatase. (B) Methylene blue stains all cells and is used to count the total colony number. (C) Results plotted as % AP+/MB+ demonstrates assay consistency among three users.
ES Cell FBS Qualified supports pluripotency in mESCs
Figure 6. Phase contrast and fluorescent images of mESC colonies. mESCs grown in media supplemented with ES Cell Qualified FBS exhibit the morphological traits of well-defined refractive edges, tight colonies with small cells, and few differentiated cells. These cells also exhibit pluripotency via Alkaline Phosphatase (AP) Live Stain and the expression of Oct4, and Sox2.