THE JOURNAL OF TROPICAL LIFE SCIENCE OPEN ACCESS Freely available online VOL. 6, NO. 3, pp. 196 - 204, September, 2016 Submitted May 2016; Revised August 2016; Accepted September 2016 Agarose Coated Culture Plate in Tumorsphere Culture of Cervical Cancer Cell Line HeLa: An Alternative to Non Adhesive Culture Plate I Putu Juniartha 1 *, Muhammad Rasjad Indra 2 , Hidayat Sujuti 3 , Diana Lyrawati 4 , Tatit Nurseta 5,6 1 Department of Biomedical Sciences, Faculty of Medicine, Brawijaya University, Malang, Indonesia 2 Department of Physiology and Molecular, Faculty of Medicine, Brawijaya University, Malang, Indonesia 3 Department of Biochemistry and Biomolecular, Faculty of Medicine, Brawijaya University, Malang, Indonesia 4 Department of Pharmacy, Department of Biomedical Sciences, Faculty of Medicine, Brawijaya University, Malang, Indonesia 5 Department of Obstetry and Gynecology, Faculty of Medicine, Brawijaya University, Malang, Indonesia 6 Department of Obstetry and Gynecology, dr. Saiful Anwar Public Hospital, Malang, Indonesia ABSTRACT Cervical cancer recurs in 90% cases and linked to cancer stem cells that able to self-renew and responsible for recurrence, metastasis, and mortality of cancer. Isolation and identification of cancer stem cells using serum-free medium needs expensive growth factors and consume time. This study try to grow tumor sphere using culture plate coated with 1% agarose as an efficient and economical alternative to non-adhesive culture plate. HeLa cell line was grew in culture plate coated with 1% agarose and non-adhesive culture plate using similar medium and culture condition. Tumor spheres morphology was observed and the colonies were counted in 7 days followed by single cell assay. Tumor spheres then counted for CD133+, CD34+, and Sox2 expression using flowcytometry. Culture plate coated with 1% agarose can be used as an economic and efficient alternative to culture tumor sphere. Using culture plate coated with 1% agarose, the tumor spheres formed in 7 days with similar morphology to non-adhesive culture plate. Tumorsphere had three dimensional – sphere shape that tightly attached, colonized, and overlapped. The tumor sphere colony counts of two plates were statistically have no significant difference (p=0,667). Single cell assay of a tumor sphere shows that it can grow new tumor spheres with similar morphology. The tumor sphere from culture plate coated with 1% agarose express CD133+ and CD34+ as much as 8.78% ± 2.14 and Sox2 as much as 35.30% ± 23.82 whereas tumor sphere from non-adhesive culture plate express CD133+ and CD34+ as much as 62.36% ± 1.06 and Sox2 as much as 98.86% ± 0.56 (p = 0000). Keywords: Cancer stem cell, tumorsphere, agarose, non-adhesive culture plate, Sox2 Cervical cancer is the second most common cancer in women after breast cancer. In every year, approxi- mately 500.000 women diagnosed with invasive cervical cancer and half of them die [1, 2]. Recurrent cervical cancer still remains a problem that requires proper treatment. Recurrency happened in almost 90% cases in 2 years after therapy and 20 – 70% are in the early stage and local advanced stage [3]. Until now, cervical cancer is treated by chemotherapy, surgery, and genetic based therapy that targets tumor oncogene and gene suppressor, but the cancer still re- curs [4]. Recent studies shows that a tumor grew from a small population in tumor cells called cancer stem cell that have several properties such as self-renewal, chemoresistance, and able to differentiate into cancer with tumorigenic properties. These properties were considered to take part in cancer recurrence, metasta- sis, and high mortality [5, 6, 7, 8]. Several study had been done to identify cancer stem cell in cervical cancer. Feng et al., (2009) successfully JTLS | J. Trop. Life. Science 196 Volume 6 | Number 3 | September | 2016 INTRODUCTION *Corresponding author: I Putu Juniartha Department of Biomedical Science Faculty of Medicine, Brawijaya University Jalan Veteran, Malang 65145, Indonesia Email: koasputu@gmail.com doi: 10.11594/jtls.06.03.11 How to cite: Juniartha IP, Indra MR, Sujuti H, Lyrawati D, Nurseta T (2016) Agarose Coated Culture Plate in Tumorsphere Culture of Cervical Cancer Cell Line HeLa: An Alternative to Non Adhesive Culture Plate. J. Trop. Life. Science 6 (3): 196 – 204.