title: Isolation, culturing and characterization of feeder-independent amniotic fluid stem cells in buffalo (Bubalus bubalis).
abstract: Heterogeneous amniotic fluid contains various cell types. The aim of this study was to characterize and differentiate some of the key stemness attributes of the amniotic fluid-derived cells in buffalo (Bubalus bubalis). The amniotic fluid (AF) cells were cultured without feeder cells, in DMEM containing 15% FBS, 1% non-essential amino acids, 1% penicillin/streptomycin/ampicillin, 1% vitamin solution, and 1% l-glutamine in 5% CO(2) in humidified air at 38.5±0.5 °C. After 6 days of culture different types of cells viz., star shaped (62.7%), spherical without nucleus (1.9%), spherical with nucleus (26.4%), pentagonal (0.4%), and free floating/rounded cells (8.3%) were observed. Most of the cells started anchorage-dependent growth after day 7 of the culture. Expression of alkaline phosphatase (AP) and Oct-4, Nestin and FGF-5 were observed from the AF cells at different passages. Using species-specific primers, a PCR amplicon of 200, 296 and 210 bp were observed for Oct-4, Nestin and FGF-5, respectively. The cells were found to have a normal karyotype at different passages. These results may contribute towards establishing non-embryonic pluripotent stem cells for various therapeutic and reproductive biotechnological applications in the species.
authors: Department of Biotechnology, Kurukshetra University, Kurukshetra 136 119, India.
authors: Kapil
authors: K
authors: Dev
authors: Sanjeev K
authors: SK
authors: Gautam
authors: Shiv K
authors: SK
authors: Giri
authors: Anil
authors: A
authors: Kumar
authors: Anita
authors: A
authors: Yadav
authors: Vinod
authors: V
authors: Verma
authors: Pushpander
authors: P
authors: Kumar
authors: Birbal
authors: B
authors: Singh
mesh_terms: D000653:Amniotic Fluid / Q000166:cytology*; D000818:Animals; D002020:Buffaloes / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods / Q000662:veterinary*; D002454:Cell Differentiation; D002455:Cell Division; D061252:Feeder Cells; D005260:Female; D059785:Karyotype; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000502:physiology*; D011247:Pregnancy