Collection Protocol of Fetal and Foreskin-derived Fibroblasts
GUIDELINE
- Embryonic fibroblasts, the major cellular component of sparse connective tissue, are differentiated from mesenchymal cells during the embryonic period.
- Fibroblasts are large, well-defined, mostly spindle-shaped or star-shaped flat structures with regular ovoid nuclei and large and obvious nucleoli. Fibroblasts have high functional activity, weak basophilic cytoplasm, and obvious protein synthesis and secretion activities.
METHODS
Fetal fibroblasts
- Take about 3 cm2 of aborted fetal abdominal skin or foreskin, place it in a 35 mm Petri dish containing DPBS, and cut it into 1 mm2.
- The tissue pieces are collected and transferred to a 15 mL centrifuge tube, and centrifuged at 1000 r/min for 3 min at room temperature.
- Discard the supernatant, resuspend the tissue blocks with cell base culture medium, and transfer to a T25 culture flask, trying to evenly distribute the tissue blocks, while carefully aspirating the culture medium around the tissue blocks.
- Invert the culture flask overnight, turn it over the next day, and add 1.5 mL of cell base culture medium.
- The culture medium is replaced with a fresh culture medium every other day, and cells are seen crawling out around the tissue block for about 7 days.
- Passage with 0.25% trypsin.
Foreskin-derived fibroblasts
- Foreskin tissue is taken and rinsed with DPBS containing 2% P/S.
- The foreskin tissues are well cut with ophthalmic scissors, added with 0.25% trypsin, and digested until the tissues are in a mucous flocculent state.
- The digestion is terminated by adding an equal volume of fibroblast culture medium (DMEM medium, 10% FBS, 1% NEAA, 1% P/S).
- The cells are blown and mixed, centrifuged and resuspended, then inoculated in culture flasks and incubated in the incubator.
- 1-3 generations of foreskin fibroblasts are selected as the donor cells for iPSCs.
Creative Bioarray Relevant Recommendations
- We provide different types of fibroblasts, including but not limited to Immortalized Human Foreskin Fibroblasts, Immortalized Human Intestinal Fibroblasts, Immortalized Human Neonatal Fibroblasts-hTERT, and others.
NOTES
The skin and foreskin of the aborted fetus are taken from the hospital. Signed informed consent is required.