Obtaining multipotent amnion-derived stem cell (ADSC) from amniotic membrane tissue without enzymatic digestion
View Patent ↗The present invention relates to stem cells obtained from the amnion and their methods of obtaining and culturing. The present invention further relates to compositions comprising amnion-derived stems cells (ADSCs) and to methods of using ADSCs.
1. A method for obtaining a multipotent amnion-derived stem cell (ADSC) comprising:
a. separating an amniotic membrane tissue sample from chorion of a mammalian embryo;
b. culturing the amniotic membrane tissue sample in culture media without any enzymes or reagents to digest the amniotic membrane tissue sample;
c. preparing a single-cell culture of adherent ADSCs isolated from the amniotic membrane tissue sample;
d. culturing the ADSCs; and
e. obtaining or isolating the ADSCs.
2. The method of claim 1 , wherein the amniotic membrane tissue sample is washed and fragmented after step a. and before step b.
3. The method of claim 1 , wherein the amniotic membrane tissue sample is cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 20% fetal bovine serum (FBS).
4. A method of cryopreserving an amniotic membrane tissue sample comprising:
a. washing an amniotic membrane tissue sample;
b. suspending the amniotic membrane tissue sample in a cryopreservation medium consisting essentially of 60% Dulbecco's Modified Eagles Media (DMEM)/30% Fetal Bovine Serum (FBS) or 30% Liforcel serum substitute/10% Dimethyl Sulfoxide (DMSO); and
c. storing the tissue sample at a temperature below about −80° C.
5. A method of cryopreserving ADSCs comprising:
a. washing a population of ADSCs;
b. suspending the population of ADSCs in a cryopreservation medium comprising 60% Dulbecco's Modified Eagles Media (DMEM)/30% Fetal Bovine Serum (FBS) or 30% Liforcel serum substitute/10% Dimethyl Sulfoxide (DMSO); and
c. storing the tissue sample at a temperature below about −80° C.