The total protein in each batch of AMEED was assessed
using a standard Bradford protein assay. Briefly, 20 µl of
each sample and a diluted standard that contained 10 µg/
µl .-globulin were added to the wells of a 96-well plate
(in duplicate), followed by the addition of 500 µl Bradford
buffer (5000006, Bio-Rad Laboratories, Inc., Hercules,
CA, USA) to each well and mixed. The optical density
at 595 nm was then measured using a spectrophotometer
(Multiskan Spectrum, Thermo Fisher Scientific Oy,
Vantaa, Finland).
The concentrations of EGF, KGF, hepatocyte growth
factor (HGF), and interleukin-1 receptor antagonist (IL1RA),
as important amniotic membrane proteins necessary
for epithelial regeneration (14 (link)), were assessed using
commercially available enzyme-linked immunosorbent
assay (ELISA) kits (Catalogue no.: DEG00, DKG00,
DHG00, and DRA00B, R & D Systems Inc., Minneapolis,
MN, USA) according to the manufacturer’s protocols.
Four batches of AMEED were used for this growth factor
analysis. The stability of the growth factors was tested
after one month to one year of storage at -70°C, after 7
days of storage in a refrigerator (2-8°C), and after 2 days
of storage at room temperature.
using a standard Bradford protein assay. Briefly, 20 µl of
each sample and a diluted standard that contained 10 µg/
µl .-globulin were added to the wells of a 96-well plate
(in duplicate), followed by the addition of 500 µl Bradford
buffer (5000006, Bio-Rad Laboratories, Inc., Hercules,
CA, USA) to each well and mixed. The optical density
at 595 nm was then measured using a spectrophotometer
(Multiskan Spectrum, Thermo Fisher Scientific Oy,
Vantaa, Finland).
The concentrations of EGF, KGF, hepatocyte growth
factor (HGF), and interleukin-1 receptor antagonist (IL1RA),
as important amniotic membrane proteins necessary
for epithelial regeneration (14 (link)), were assessed using
commercially available enzyme-linked immunosorbent
assay (ELISA) kits (Catalogue no.: DEG00, DKG00,
DHG00, and DRA00B, R & D Systems Inc., Minneapolis,
MN, USA) according to the manufacturer’s protocols.
Four batches of AMEED were used for this growth factor
analysis. The stability of the growth factors was tested
after one month to one year of storage at -70°C, after 7
days of storage in a refrigerator (2-8°C), and after 2 days
of storage at room temperature.