Nimotuzumab is an anticancer agent which belongs to the inhibitor group of Epidermal Growth Factor Receptor (EGFR). This monoclonal antibody has a relatively high molecular weight which makes slow penetration on tumor cell, as concequence, it is less attractive in imaging kinetics, and potentially elicits antibodies respons. Therefore in this study nimotuzumab was fragmented to form bivalent antibody [F(ab')2] and then labeled with 125I to form 125I-F(ab')2-nimotuzumab which can be used further as a precursor for preparing 125I-F(ab')2-nimotuzumab-NLS (NLS = nuclear localizing sequences) radiopharmaceutical for radioimmunotherapy. The aims of this study were to obtain characteristics of 125I-F(ab')2-nimotuzumab by comparing with the 125I labeled-intact nimotuzumab (125I-nimotuzumab). This study was initiated by purifying nimotuzumab by mean of dialysis. The purified nimotuzumab was then fragmented by using pepsin. The F(ab')2-nimotuzumab formed was then purified from its by-products which formed in fragmentation process by using a PD-10 column (consisted Sephadex G25). The intact nimotuzumab and its F(ab')2 fragment were then labeled with the 125I to form 125I-nimotuzumab and 125I-F(ab')2-nimotuzumab. The radiochemical purity are 98.27 % and 93.24 %, respectively. Stability test results show that, both of 125I-nimotuzumab and 125I-F(ab')2-nimotuzumab more stable at 4 °C than at room temperature storage and 37 °C.