Human dental pulp tissue was obtained from eight healthy volunteers aged 22 to 40 years undergoing routine extraction of third molars and premolars for orthodontic reasons. The isolation and expansion of DPSCs were performed as previously described [24 (link), 37 (link)]. Briefly, the enzymatically isolated cells were cultured in XFM (PRIME-XV MSC Expansion XSFM; Irvine Scientific, Santa Ana, CA, USA) in a humidified incubator at 37 °C and 4.7% CO2. Upon reaching 80% confluence, the cells were dissociated using 0.25% trypsin–0.02% ethylenediaminetetraacetic acid (EDTA) buffer and then subcultured at a cell density of 5 × 103 cells/cm2 in 60-mm culture dishes coated with the substrates as determined by the following pilot experiment. All experiments were performed with cell passages P3–P5. To compensate for donor variability, all experiments were independently repeated using cells obtained from at least four individuals, with the exception of the transmission electron microscope (TEM) analysis.
Prime xv msc expansion xsfm
Manufactured by Irvine Scientific
Sourced in United States
PRIME-XV MSC Expansion XSFM is a serum-free, xeno-free medium designed for the expansion of human mesenchymal stem cells (MSCs) in cell culture.
Automatically generated - may contain errors
Lab products found in correlation
5 protocols using prime xv msc expansion xsfm
1
Isolation and Expansion of DPSCs
2
Induced Mesenchymal Stem Cell Generation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
iMSCs were generated by inducing hiPSCs into neural crest cells (NCCs), and then into iMSCs (Kamiya et al., 2022 (link)), which were then maintained in a plastic dish coated with Fibronectin (Chemicon, FC010-10 MG) and fed with PRIME-XV®MSC Expansion XSFM (IrvineScientific, 91149). Analyses were performed during passages 3—5.
3
Preparation of Cell-Sheet Derived MSCs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
C-MSCs were prepared as reported previously with minor modifications [34 (link)]. Briefly, human bone marrow MSCs provided by RIKEN BIORESOURCE CENTER (Tsukuba, Ibaraki, Japan) were seeded at a density of 1.0 × 105 cells/well in 48-well plates (Corning, Corning, NY, USA) and cultured with Prime-XV MSC expansion XSFM (Irvine Scientific, Santa Ana, CA, USA) for 4 days. To obtain C-MSCs, confluent cells that had formed on the cellular sheet, consisting of the ECM produced by MSCs themselves, were scratched by using a micropipette tip and then torn off. The C-MSCs detached from the bottom of the plate in a sheet shape were transferred to a 24-well ultra-low-binding plate (Corning) and rolled up to make a round clump of cells. The cell clumps were maintained in MSCgo Osteogenic-SF, XF medium (Biological Industries, Beit Haemek, Israel) for 3 days (Figure 1 A).
4
Chondrogenic Differentiation of C-MSCs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Human C-MSCs were generated with XF condition as previously reported [17 (link)]. Briefly, passage number 5 (population doubling number is approximately 7 to 8) of commercially available human bone marrow MSCs (26 years old male donor; LONZA, Basel, Switzerland) were seeded at a density of 1.0 × 105 cells/well in 48-well plates (Corning, Corning, NY, USA) and maintained in Prime-XV® MSC expansion XSFM (GM) (Irvine Scientific, Santa Ana, CA, USA) for 4 days. To obtain C-MSCs, confluent cells that had formed on the cellular sheet, consisting of the ECM produced by MSCs themselves, were scratched by using a micropipette tip and then torn off. The MSC/ECM complex was detached from the bottom of the plate in a sheet shape and transferred to a 24-well ultra-low-binding plate (Corning). Then, the cellular sheet rolled up to make a round clump of cells, so called C-MSCs. The cell clumps were cultured in GM or MSCgo™ Chondrogenic XF medium (CIM) (Biological Industries, Beit Haemek, Israel) for 3, 5, 7, 10, 13, and 15 days.
5
Dental Pulp Stem Cell Expansion and CFU Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
DPSCs were expanded under two different culture conditions: XSF (PRIME-XV® MSC expansion XSFM, Irvine Scientific, USA) or MEM-α medium containing FBS (10%). Briefly, when DPSCs from passage 1 reached 80-90% confluence, the cells were enzymatically detached, counted, and re-seeded in XSF or FBS medium at a cell density of 30,000 cells/25 cm 2 flask and cultured for 1 week without medium change. Subsequently, the cells were counted with a hemocytometer and replated every week through to passage 6. Population doublings (PDs) were calculated using the following Eq. 1:
DPSCs from passage 2 and passage 6 were used in the CFU assay. Briefly, 300 DPSCs were seeded to a 25-cm 2 flask and cultured in XSF or FBS medium for 2 weeks without medium change. At the end of culture, the DPSCs were fixed and stained with 0.1% (w/v) toluidine blue prepared in 2% (w/v) paraformaldehyde for 1 h, and subsequently rinsed with distilled water. Colonies (≥ 50 cells) were then counted and scored as a CFU. The experiments were performed using samples three different donors (N = 3; n = 3).
DPSCs from passage 2 and passage 6 were used in the CFU assay. Briefly, 300 DPSCs were seeded to a 25-cm 2 flask and cultured in XSF or FBS medium for 2 weeks without medium change. At the end of culture, the DPSCs were fixed and stained with 0.1% (w/v) toluidine blue prepared in 2% (w/v) paraformaldehyde for 1 h, and subsequently rinsed with distilled water. Colonies (≥ 50 cells) were then counted and scored as a CFU. The experiments were performed using samples three different donors (N = 3; n = 3).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!