Pt 4124

Manufactured by Lonza

The PT-4124 is a laboratory equipment product. It is a device designed for use in a laboratory setting. The core function of the PT-4124 is to perform specific laboratory tasks, but the details of its intended use are not available.

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Lab products found in correlation

Pt 3003, by Lonza (3 mentions) 4000b dmi, by Leica (1 mentions) Alcian blue, by Merck Group (1 mentions) Tgf β3, by Lonza (1 mentions) Dexamethasone (dex), by Merck Group (1 mentions) Insulin, by Merck Group (1 mentions) Indomethacin, by Merck Group (1 mentions) β glycerophosphate, by Merck Group (1 mentions) Ascorbic acid, by Merck Group (1 mentions) Alizarin red s, by Merck Group (1 mentions) Trypsin edta, by Fujifilm (1 mentions) Mubmx 01001, by Cyagen (1 mentions) Muxmx 90011, by Cyagen (1 mentions) Tri reagent, by Molecular Research Center (1 mentions) Biomasher, by Nippi (1 mentions)

4 protocols using pt 4124

Multilineage Differentiation of MSCs

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Cells were grown from cryopreserved stocks to confluence in twelve-well-plates and then subjected to specific differentiating conditions using the osteogenic differentiation medium PT-4120, and chondrogenic medium PT-3003 supplemented with 10 ng/mL TGFβ3, PT-4124 (all from Lonza) under conditions described by the manufacturer.
Multilineage differentiation potential of MSCs was assessed by histochemical staining and phase contrast microscopy (Leica 4000b DMI, Leica Microsystems GmbH. Germany).
The degree of mineralization in osteogenic cultures was assessed staining with 2% Alizarin Red S (Sigma-Aldrich-Aldrich). Briefly, cells were washed three times with phosphate buffered saline (PBS) pH 4.2 and fixed 20 minutes with 4% paraformaldehyde. Fixed cells were washed and stained and washed again to remove excess of stain. Similarly, chondrogenic potential was evaluated measuring production ECM proteoglycan produced by chondrocytes after staining for 20 minutes with 1% Alcian blue (Sigma-Aldrich).

Tornero-Esteban P., Peralta-Sastre A., Herranz E., Rodríguez-Rodríguez L., Mucientes A., Abásolo L., Marco F., Fernández-Gutiérrez B, & Lamas J.R. (2015). Altered Expression of Wnt Signaling Pathway Components in Osteogenesis of Mesenchymal Stem Cells in Osteoarthritis Patients. PLoS ONE, 10(9), e0137170.

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Multilineage Differentiation of BMSCs

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To induce osteogenic differentiation, we cultured BMSCs in α-MEM supplemented with 20% heat-inactivated FBS, 10 nM dexamethasone (D1759), 100 mM ascorbic acid (49752), and 10 mM β-glycerophosphate (G9422, all from Millipore Sigma). For evaluation of osteogenesis, cells were fixed in 4% paraformaldehyde (PFA), washed twice with PBS, and stained with 2% Alizarin red S solution (pH 4.3; A5533, Millipore Sigma). Adipogenic differentiation was induced with Dulbecco’s modified Eagle’s medium (high glucose) supplemented with 10% heat-inactivated FBS, 0.5 mM indomethacin (I7378, Millipore Sigma), 1 μM dexamethasone, and 10 μg/mL insulin (91077C, Millipore Sigma). The differentiation medium was changed every 2–3 days. For evaluation of adipogenesis, cells were fixed in 4% PFA and stained with filtered oil red O solution for 15 min at 37°C. After washing with 60% isopropanol, cells were either imaged or oil red O extracted with 100% isopropanol and quantified by the absorption of 500 nm with a spectrophotometer. To induce chondrogenic differentiation, we cultured BMSCs in MSC chondrogenic differentiation medium (PT-3003, Lonza, Allendale, NJ) supplemented with 10 ng/mL TGFβ3 (PT-4124, Lonza) according to the manufacturer's instructions. For evaluation of chondrogenesis, cell aggregates were cryosectioned, fixed in 4% PFA, stained with toluidine blue O solution (pH 4.5), and imaged.

Hu T., Kitano A., Luu V., Dawson B., Hoegenauer K.A., Lee B.H, & Nakada D. (2019). Bmi1 Suppresses Adipogenesis in the Hematopoietic Stem Cell Niche. Stem Cell Reports, 13(3), 545-558.

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Chondrogenic Differentiation of Mouse MSCs

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Mouse MSCs were purchased from cyagen (MUBMX-01001) and cultured in a complete growth medium (cyagen, MUXMX-90011). Mouse MSCs were digested in 0.5% trypsin-EDTA (Wako), and trypsin was inactivated with 2× volume of expansion medium. Dissociated cells were centrifuged at 150×g for 5 min, and the supernatant was aspirated. Subsequently, cells were washed in incomplete chondrogenic induction medium (Lonza, PT-3003), and resuspended at 5 × 10⁵ cells/mL in complete chondrogenic medium: incomplete chondrogenic induction medium and 10 ng/mL rhTGF-β3 (Lonza, PT-4124). In total, 500 µL of the above cell mixture was dispensed into 15-mL conical tubes and centrifuged at 200 ×g for 5 min. Pellets were cultured at 37 °C in 5% CO₂ for 21 days with medium exchanged every 3 days. Each pellet was homogenized in TRI Reagent (Molecular Research Center) using a BioMasher (Nippi). Subsequent steps were performed using the manufacturer’s protocol.

Ito Y., Matsuzaki T., Ayabe F., Mokuda S., Kurimoto R., Matsushima T., Tabata Y., Inotsume M., Tsutsumi H., Liu L., Shinohara M., Tanaka Y., Nakamichi R., Nishida K., Lotz M.K, & Asahara H. (2021). Both microRNA-455-5p and -3p repress hypoxia-inducible factor-2α expression and coordinately regulate cartilage homeostasis. Nature Communications, 12, 4148.

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Chondrogenic Differentiation of Pericytes

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Pericytes were aliquoted into 15 mL polypropylene culture tubes at 1 × 10⁶ cells/tube. The cells were centrifuged at 150× g for 5 min at room temperature. The centrifuged cells were incubated with chondrogenic induction medium (Lonza, #PT-3925) supplemented with dexamethasone, ascorbate, ITS (insulin, transferrin and selenium), GA-1000, sodium pyruvate, proline, L-glutamine (Lonza, #PT-4121), 5 ng/µL TGF-β3 (Lonza, #PT-4124), and 100 ng/µL BMP-6 at 37 °C for 4 weeks. The medium was changed every 2–3 days. After 4 weeks of culture, half of the pellets were fixed with 4% paraformaldehyde, embedded in paraffin, and cut into 5 µm sections for alcian blue staining. The other half of the pellets was used for quantitative PCR analyses.

Supakul S., Yao K., Ochi H., Shimada T., Hashimoto K., Sunamura S., Mabuchi Y., Tanaka M., Akazawa C., Nakamura T., Okawa A., Takeda S, & Sato S. (2019). Pericytes as a Source of Osteogenic Cells in Bone Fracture Healing. International Journal of Molecular Sciences, 20(5), 1079.

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