New Methods for Establishment of Lymphoblastoid Cell Lines

Abstract

Background: Human lymphocytes can be isolated from whole blood by centrifugation using a commercially available high-density medium. This allows a single step gradient separation of blood, which yields the mononuclear cell ,the lymphocyte can be cultured specifically and will outgrow the others, eventually resulting in highly enriched population.Objective: this series of experiments were done as a trial to establish a new method for preparation of lymphocytes cell line.Methods: the same protocols were applied to all individuals, they were separated in to two groups:- Group 1:- one ml of heparinized blood were cultured in tissue culture flask containing 9 ml of stimulation medium and incubated in tissue culture flasks at 37ºC (4) . The flasks were subcultured at 37 OC in CO2 incubator every 3 days.Group 2:- Mononuclear cells were separated from whole blood, washed, counted, assessed for viability, then 1 ml of 2x106 cells were cultured in tissue culture flask containing stimulation medium and. The flask was incubated at 37 OC in CO2 incubator for 3 days. Another 1ml of mononuclear cells was cultured in tissue culture flask containing 1ml of growth medium, the flasks were incubated at 37 OC in CO2 incubator for 3 days.For both groups the cell cultures were propagated and maintained. Results: After initiation of cell culture for both groups, the mononuclear cell cultures were maintained up for 3 weeks, pure rich mononuclear cells were obtained and seen under inverted microscope.Conclusion: a new method for establishment of lymphoblast cell line were developed by cultivation of lymphocyte from whole blood . pure rich lymphocytes culture were obtained and maintained