KULTUR IN VITRO SEL-SEL OTAK BESAR (CEREBRUM) ANAK TIKUS
(In Vitro Culture of Newborn Rat Cerebrum Cells)
VIVIT RIYACUMALA, ITA DJUWITA
ABSTRAK
Telah dilakukan penelitian kultur in vitro sel-sel otak besar anak tikus (Sprague Dawley) umur tiga hari dalam Dulbecco’s Modified Eagle Medium (DMEM) yang mengandung Fetal Bovine Serum (FBS) 10%, dan gentamisin 50 μg/ml dengan dan tanpa penambahan insulin 5 μg/ml, transferin 10 μg/ml, selenium 5 μg/ml (ITS). Kultur dilakukan dalam inkubator CO2 5% dan suhu 37oC selama 11 hari. Identifikasi dilakukan terhadap jenis sel yang tumbuh berdasarkan pengamatan morfologi, kemampuan pertumbuhan sel (tingkat proliferasi dan population doubling time (PDT) serta panjang akson dendrit) berdasarkan penghitungan sel menggunakan hemositometer dan mikrometer, serta kandungan protein yang dihasilkan berdasarkan identifikasi menggunakan metode Sodium dodecyl sulfate – polyacrilamide gel electrophoresis (SDS-PAGE). Data kuantitatif dianalisis menggunakan uji statistik T-test dalam program Minitab, sedangkan data kualitatif dianalisis secara deskriptif. Hasil penelitian menunjukkan bahwa berdasarkan pengamatan morfologi terdapat dua jenis sel yaitu sel saraf yang meliputi sel saraf bipolar dan multipolar serta sel glia yang meliputi astrosit fibrous, astrosit protoplasmik, oligodendrosit, dan mikroglia. Penambahan ITS ke dalam medium kultur mampu meningkatkan tingkat proliferasi (P<0,05) dengan PDT yang lebih rendah, meningkatkan pertumbuhan panjang akson dan dendrit, serta secara kualitatif produksi protein (berukuran antara 21,5–36,5 kDa) yang lebih tinggi yang diindikasikan dari intensitas pita protein.
Kata kunci: kultur sel, sel saraf, ITS, SDS-PAGE
ABSTRACT
Research has been conducted on in vitro culture of three days old rat cerebrum cells in Dulbecco’s Modified Eagle Medium (DMEM) containing 10% Fetal Bovine Serum (FBS) and 50 μg/ml gentamycin, and supplemented with and without 5 μg/ml insulin, 10 μg/ml transferin, 5 μg/ml selenium (ITS). Culture was done in 5% CO2 incubator at 37oC for 11 days. Identification was done on types of cell based on morphological observation, growth capacity (cell proliferation, population doubling time/ PDT and length of axon and dendrit) based on calculation of the number of cells using hemocytometer and micrometer, and the protein produced by the cerebrum cells in the culture medium using sodium dodecyl sulfate–polyacrilamide gel electrophoresis (SDS-PAGE) method. Quantitative data were analyzed using statistical T-test on Minitab program. The results showed that based on the morphological observations, there are two types of cells including nerve cells of bipolar and multipolar neurons and glial cells including the fibrous astrocyte, protoplasmic astrocyte, oligodendrocyte, and microglia. Supplementation of ITS into the culture medium could increase the cells proliferation rate (P<0.05) with lower PDT, axon and dendrit lenght growth and quantitatively the 21.5–36.5 kDa protein production as indicated by the intensity of the protein band.
Keywords: cell culture, nerve cell, ITS, SDS-PAGE