Neuromics' FGF-2 for Expanding Human Cells

FGF-2 and Fibrochondrocytes

FGF-2 of FGF-basic is an important a growth factor for many cell-based assays. Neuromics' has rock solid FGFs for supplementing media used to grow cells. Check out publications referencing use of FGF-2.

Here's the latest publication: Yan Liang, Enaam Idrees, Stephen H. J. Andrews, Kirollos Labib, Alexander Szojka, Melanie Kunze, Andrea D. Burbank, Aillette Mulet-Sierra, Nadr M. Jomha & Adetola B. Adesida. Plasticity of Human Meniscus Fibrochondrocytes: A Study on Effects of Mitotic Divisions and Oxygen Tension. Scientific Reports 7, Article number: 12148 (2017) doi:10.1038/s41598-017-12096-x. ...Thereafter the number of viable MFCs were counted using a haemacytometer after trypan blue staining. MFCs were plated at 104 cells/cm2 and cultured in the standard medium described above supplemented with FGF-2 (5 ng/mL; Neuromics, MN, USA, Catalog#: PR80001) and TGFβ1 (1 ng/mL; ProSpec, NJ, USA, Catalog#: cyt-716) under normal oxygen tension (21% O2) at 37 °C in a humidified incubator...

Images: Immunofluorescence analysis of collagen I and collagen II in pellets derived from T1F2-expanded MFCs of four passages after 21 days chondrogenic stimulation under NRX or HYP from one representative donor (male, 20 years old). Blue (DAPI): cells, Red (Texas Red): collagen I, Green (FITC): collagen II. (A) Pellets cultured under NRX, (B) Pellets cultured under HYP from four passages. Scale bar: 100 µm

Our ISO-Kine FGF-2 is especially potent as it virtually endotoxin free.

Neuromics' ISOKine bFGF in 3-D Cultures

Works Well in Perfusion Models

Our ISOKine^TM FGF is produced in the endosperm tissue of barley grain (Hordeum vulgare), that exhibits up to 50 times less protease activity than E.coli or mammalian cells. Barley seed is void of any human or animal viral contaminants that could jeopardize your cell culture.

It is a proven solution for all cell cultures and starts at the low price of  65 USD/10 ug.

Here's a reference of its use in 3-D Cultures: Tom Kamperman, Sieger Henke, Claas Willem Visser, Marcel Karperien, Jeroen Leijten. Centering Single Cells in Microgels via Delayed Crosslinking Supports Long-Term 3D Culture by Preventing Cell Escape. DOI: 10.1002/smll.201603711.

Figure: Delayed on-chip crosslinking enables centering of single cells in microgels. a) Fluorescence confocal imaging confirmed that delayed enzymatic crosslinking enabled centering of single MSCs in Dex-TA microgels. b) On average, cell-laden microgels were only 9 µm larger than the encapsulated MSCs, effectively resulting in 3D hydrogel coatings of less than 5 µm. c) A standard microfluidic droplet generator was connected to the H2O2 diffusion-based crosslinking chip. The position of cells (white arrows) in non-crosslinking microgel precursor droplets was analyzed d) immediately after droplet generation (t1), at the start of the crosslinking chip (t2), and e) at the end of the crosslinking chip (t3). f) Cell positions within microgels produced using conventional microfluidic encapsulation systems (i.e., with coupled emulsification and gelation) are indicated with gray (i.e., references) and red (i.e., this work) data points. Cell positions within gel precursor droplets along the modular microfluidic setup are indicated with blue data points. Cell positions within delayed enzymatically crosslinked microgels are indicated with green data points. g) Cell position analyses of various combinations of distinct hydrogel materials (i.e., Dex-TA, Dex-HA-TA, PEGDA), cell types (i.e., MIN6, MSC), and crosslinking methods (i.e., enzyme-based and photo-crosslinking), revealed that delayed crosslinking consistently resulted in significantly increased cell-centering as compared to the conventional encapsulation approach where emulsification and gelation are coupled.

If you are looking for competitively priced, animal free and potent growth factors. check out our ISOkines.

Culturing Stem Cells in 3-D

Requires Potent Media + Supplements

Neuromics is responding to the many challenges our clients face in building 3-D, in-vivo like, cell- based assays. We do this by offering the most potent Media plus Supplements like FGFS.

Here's a protocol for single cell 3-D assays using hMSCs and Hydrogels. It features use of our ISOKine^TM FGF

Images: Cell-centering in cytocompatible microgels enables long-term single-cell 3D culture by preventing cell escape. a) Qualification of Dex-TA microgel crosslinking as a function of the microemulsion flow rate (Qemulsion) and concentration of the H2O2 feed ([H2O2]feed). Blue, green, and red indicate incomplete crosslinking, complete crosslinking, and H2O2 excess, respectively. b,c) Amplex Red assay to quantify the concentration of residual H2O2 ([H2O2]emulsion) in Dex-TA microgel precursor droplets and crosslinked microgels after their retrieval from the diffusion-based crosslinking platform. d) The microencapsulation procedure had no detrimental effect on short-term cell survival. e) Delayed crosslinking resulted in 4 ± 1% cell escape after 7 d of in vitro culture, as compared to 27 ± 5% cell escape when using coupled emulsification and gelation. f) The number of encapsulated cells per microgel tightly followed the Poisson distribution and remained similar throughout long-term (28 d) of in vitro culture, which confirmed that cell centering prevents cell escape. g–i) MSCs encapsulated in delayed enzymatically crosslinked microgels remained viable and metabolically active throughout 28 d of in vitro culture. j) Positive Oil Red O and k) Alizarin Red staining confirmed that l) more than 60% of the microencapsulated MSCs could differentiate into the adipogenic and osteogenic lineage, respectively. Black scale bars: 50 µm, white scale bars: 5 µm. DOI: 10.1002/smll.20160371.

Protocol for Cell Isolation and Expansion: Human MSCs were isolated from fresh bone marrow samples and cultured as previously described. The use of patient material was approved by the local ethical committee of the Medisch Spectrum Twente and informed written consent was obtained for all samples. In short, nucleated cells in the bone marrow aspirates were counted, seeded in tissue culture flasks at a density of 500 000 cells cm−2, and cultured in MSC proliferation medium, consisting of 10% FBS, 100 U mL−1 penicillin, 100 mg mL−1 streptomycin, 1% GlutaMAX, 0.2 × 10−3 m ascorbic acid, and 1 ng mL−1 bFGF (added fresh) in αMEM. Mouse insulinoma MIN6-B1 cells (provided by Dr. P. Halban, University Medical Center, Geneva, Switzerland) were cultured in MIN6 proliferation medium, consisting of 10% (v/v) FBS, 100 U mL−1 penicillin, and 100 mg mL−1 streptomycin, and 71 × 10−6 m 2-mercaptoethanol (added fresh) in DMEM. When cells reached near confluence, the cells were detached using 0.25% Trypsin-EDTA at 37 °C and subsequently subcultured or used for experimentation.

I am at your beck and call to answer questions on our Cell Based Assay Solution. Pete Shuster-CEO and Owner, direct phone: (612) 801-1007 or pshuster@neuromics.com.