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Title: Vitrification of water buffalo (Bubalus bubalis) fetal skin fibroblast cells
Authors: Meena, C R
Das, S K
Keywords: buffalo
fetal skin fibroblasts
Issue Date: Apr-2006
Publisher: CSIR
IPC CodeInt. Cl.8 A23B4/09
Abstract: A method was developed for the cryopreservation of buffalo fetal skin fibroblast cells by vitrification. Skin cells were isolated from 1-2-month-old fetuses obtained from an abattoir, by enzymatic digestion (0.5% w/v trypsin + 0.05% w/v collagenase in DPBS) for 15-20 min. They were washed 5-6 times with DPBS and then once with RPMI-1640 + 10% FBS by centrifugation at 600 rpm. The cells were then cultured in the same medium in a CO2 incubator (5% CO2 in air) at 38.5°C for 3 d. After 3 days, the fibroblast cells from the monolayer were harvested by treating with trypsin-versene solution. They were equilibrated and loaded into 0.25 mL French straw, containing 0.5 M sucrose solution in DPBS. The straws were sealed and precooled by keeping them in liquid nitrogen (LN2) vapour for 1-2 min, following which these were plunged into LN2. After 5 days of storage the straws were warmed quickly by transferring them to a water bath at about 20ºC for 15-20 sec. The cells were collected by centrifugation and plated in 25 cm² culture flasks with 5-6 mL of culture media and incubated at 38.5º±1ºC with 5% CO2 in humidified (95%) incubator. The fibroblast cells growth started within 3 h of culture set-up and on day 3 the monolayer formation occurred.
Page(s): 236-238
ISSN: 0975-0967 (Online); 0972-5849 (Print)
Source:IJBT Vol.05(2) [April 2006]

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