Whole cell immunization is a novel strategy that the antigen overexpression cell line can be used as the immunogen. Compared with the normal immunization strategies, this strategy can benefit from higher immunogenicity, naïve antigen conformation and avoid the complex purification process, which is specifically suitable for novel antibody discovery against membrane protein (extracellular domain). Besides, this strategy is also a suitable method to identify new biomarkers from the surface proteins of tumor cell lines, thus can contribute to the cancer diagnosis and also the development of novel therapeutic reagents.