Table 2 Summarization of all used protocols for isolation of germline stem cells from different species ovary (NS: Not Stated)
Method | Author year & reference | Species (s) | Age | Marker | OSC traits | OLC size | Culture duration | Putative OSC’s origin | Colony shape and formation duration | Superiority and limitation | Growth factors used |
|---|---|---|---|---|---|---|---|---|---|---|---|
FACS | Human Mouse (C57BL/6) | 6–8 weeks mice 20s, 30s, 40s, 50s women | COOH–DDX4 | 5–8 μm primitive germ cells genetic signature | 35–50 μm | 18 months | Bone Marrow and peripheral blood | 10–12 weeks in mouse 4–8 weeks in human | Viable and purified population | N-2 supplement, LIF EGF bFGF GDNF | |
Human | Reproductive ages and menopause | SSEA-4 | 2–4 μm and bigger cells 8 μm small, round and yellow-color | 60 μm | 21–23 days | an integral part of the ovarian surface epithelium | approximately 3 months | Homogenous population | Without growth factors | ||
Dunlop 2014. [79] | Bovine Human | NS | DDX4 | NS | NS | several month | NS | NS | NS | NS | |
OSE Scraping | Human | Reproductive ages and menopause | OSCs separated by density gradient centrifugation | Small round cells with a bubble-like structure | 95 μm | 20 days | PGCs and VSELs | NS | Simple and easy | Without growth factors | |
Human | 27–38 years women | Whole ovarian cells were cultured | 10 μm nuclear MAPK & PS1 immunoexpression | 180 μm | 5–6 days | mesenchymal Somatic cells in the tunica albuginea | NS | Simple and easy | Without growth factors | ||
Bhartiya 2011 [33] | Human Monkey Sheep Rabbit | Menopausal women with a mean age range of 46 years | Whole ovarian cells were cultured | Smaller PSCs: 1–3 μm Larger PSCs: 4–7 μm Dark, bubbly, shiny appearance Large, darkly stained nuclei with a thin rim of cytoplasm | 130 μm | 3 weeks | VSELs origin | Day 10 Flat with a well-defined margin ES cell-like colonies 3-dimensional dense floating embryoid body-like structures | Simple and easy | Without growth factors | |
MACS | Johnson 2005 [21] | Mouse | 6 and 9Â weeks | SSEA-1 | Isolated cells were used directly for RT-PCR | Bone Marrow and peripheral blood | NS | NS | NS | ||
Zou 2009 [18] | Mouse (C57BL/6) | Adult and 5 day old female | MVH | Large round or ovoid cells with little cytoplasm and spherical nuclei with slight staining, a large ratio of nuclear plasma and nuclear diameter of 12–20 μm | NS | 15 months and more for neonatal FGSCs 6 months for adults FGSCs | NS | After 7–8 passages (forming cluster) | Poor describing details Relatively low efficiency | LIF Transferrin Insulin Putrescine EGF GDNF bFGF | |
Zou 2011 [43] | CD-1 mice | Fragilis | 5-day-old female | NS | NS | NS | NS | NS | Improved Efficiency | LIF Transferrin Insulin Putrescine EGF GDNF bFGF | |
Rat | 5-day-old | Fragilis | Round with a high nuclear to cytoplasm ratio | 55–60 um | More than one year | NS | In first 3 weeks: spherical or grape-like clusters consisting of 4–8 cells | Better efficiency of gene transfer for producing transgenic rat | LIF EGF GDNF bFGF | ||
size and morphology similar to mouse FGSCs | |||||||||||
Bui 2014 [69] | Pig | Prepubertal gilts | SSEA-4 | 5–7 μm Completely round nuclei that took up almost the entire volume of the cell | >100 μm | 6 months | VSELs and epiblast-derived PGCs | Spherical colonies comprising compact clusters of small round after 1 day Dark and shiny colonies | Better culture condition for establishment and long-term maintenance of PSCs | B27 SCF | |
Using ΔPE-Oct4-Gfp transgenic mice | Pacchiarotti 2010 [48] | Mouse | 2 and 5 days old or one adult mouse | GFP which is express under Oct-4 promoter | 10–15 μm Located in OSE positive for VASA, c-Kit, SSEA-10.05 % GFP positive in adults and 1–2 % in neonata | Up to 60 μm | More than one year | They are reserved pool in the quiescent state | One week | Is not optimal for several reasons [20] | Insulin Transferrin Selenium Fibronectin |
Morphology based selection methods | Parvari 2015 [50] | Mouse | 5–7 day old | OSCs separated by colony selection | 2–4 μm As same as VSELs | Slightly bigger than OSCs | 11 days | VSELs origin | 4 days after pre-plating Small colonies were highly compact without a clear border | Easy Affordable | LIF |