KULTUR IN VITRO SEL-SEL FIBROBLAS FETUS TIKUS
HARLYSTIARINI, ITA DJUWITA
ABSTRAK
Penelitian yang dilakukan berupa kultur in vitro sel-sel fibroblas fetus tikus menggunakan Dulbeccos’ Modified Eagle Medium yang dimodifikasi dengan penambahan 3mM natrium bikarbonat, 10% fetal calf serum, dan 50 μg/ml gentamisin (mDMEM) di dalam inkubator CO2 pada suhu 37ºC. Evaluasi dilakukan terhadap tingkat proliferasi dan produksi protein setelah beberapa pasase. Pertumbuhan sel-sel fibroblas diidentifikasi melalui pengamatan morfologi. Tingkat proliferasi sel ditentukan berdasarkan population doubling time (PDT) dengan menghitung jumlah sel pada setiap pasase menggunakan hemositometer Improved Neubauer. Identifikasi protein dilakukan dengan metode Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE). Berdasarkan pengamatan morfologi, sel yang berkembang di dalam kultur in vitro adalah sel fibrosit dan sel fibroblas dan memperlihatkan peningkatan homogenitas setelah beberapa pasase. Berdasarkan nilai PDT, tingkat proliferasi sel juga meningkat seiring pasase. Sel fibroblas juga tetap mensekresikan protein hingga pasase 9 berdasarkan hasil elektroforesis. Dengan demikian dapat disimpulkan bahwa pertumbuhan dan sekresi protein dari kultur in vitro sel-sel fibroblas bisa dipertahankan sampai 9 pasase.
Kata kunci: kultur sel, fibroblas, galur sel, SDS PAGE
ABSTRACT
Research has been conducted on the in vitro culture of rat fetal fibroblast using Dulbeccos’ Modified Eagle Medium supplemented with 3mM sodium bicarbonate, 10% fetal calf serum and 50 μg/ml gentamycin (mDMEM) in CO2 incubator at 37oC. Evaluation was done on the proliferation rate and the protein production after several passages. The growth of the fibroblast was confirmed by their morphology. The proliferation rate was done based on the population doubling time (PDT) by counting the number of cells in each passage by using Improved Neubauer hemocytometer. Identification of proteins was done quatitatively using Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE). Based on morphological observation the cell grown in in vitro culture were the fibrocyte and fibroblast and showed increasing of homogenicity after several passages. Based on the PDT, the proliferation rate was increased by each passages. Fibroblast also secrete proteins remain until passage 9 based on the results of electrophoresis. It can be concluded that the growth and protein secretion of rat fetal fibroblast in in vitro culture can be maintained up to 9 passages.
Keywords: cell culture, fibroblast, cell line, SDS PAGE