**6.9 Cytotoxicity evaluation of biomaterial**

Cytotoxicity evaluation of isolated biopolymer was done on the Neuroblastoma cell line. The materials used are Cell line–SHSY-5Y, (human breast cancer cell line), Ham F-12 media, Fetal Bovine Serum (FBS), the antibiotic-antimicotic solution from Thermo scientific and MTT reagent from Sigma Aldrich, USA. Tissue culture flask, 96,6 well micro-culture plates from Eppendorf, Germany. In the method, the maintenance of cell lines, the subculturing procedure of cell lines, trypsinization, cryopreservation of cell lines was done. In MTT assay the formazan product is analyzed spectrophotometrically (540 nm) after dissolution in DMSO, the spectra of treated and untreated cells giving an estimate of the extent of cytotoxicity [13, 66].
