A Specific Human ScFv Against Malignant Melanoma Selected From A Large Human Phage Antibody LiBrary

Malignant melanoma (MM) is a kind of tumor that often occurs in skin, grows rapidly and leads to death with a high frequencies and is still on increase worldwide. The primary therapy for MM is excision surgery for MM is not sensitive to standard radiotherapy and chemotherapy. However, surgery can only be curative for original MM. Hence, experimental forms of treatment such as immunotherapy, tumor targeted therapy and genetic therapy have been studied extensively in the treatment of MM, part of which has been proved to be effective in decreasing metastasis, postponing recrudescence, and increasing survival rate of MM.Antibody targeted therapy was used early in the 1980s, in which antibodies are conjugated to chemotherapy drugs, toxins, radionuclide, enzyme or genes for targeted therapy. However,antibody targeted therapy is still on study now and has not been widely used in clinics since specific antigens of MM are very limited, so are specific antibodies. Researchers have been throwing themselves into searching antibodies of MM with high specificity and affinity. Phage display technology is the best way to solve this problem at present.Phage antibody LiBrary is constructed by displaying the whole set of antibody genes on phage surface, so it is very convenient to select specific antibodies with different antigens. Furthermore, those phage antibody LiBraries with large capacity and good diversity have the advantage of obtaining antibodies with high affinity. Phage display technology has already become one of the key techniques of antibody engineering. Antibodies against MM obtained from phage antibody LiBrary would be effective to treat MM.The LiBrary was panned with a certain kind of MM cells (LiBr) for 4 rounds. Random Clones in round NO.3 or 4 were selected to express phage antibodies. The antigen binding activities of random clones were tested by ELISA in order to select specific antibodies, which were then confirmed by DNA sequencing and immunohistochemistry. In conclusion, the enrichment of the output/input ratio was about 53 times after 4 rounds of screening, and the positive clone selected from the 80 random clones was able to bind one kind of MM cell (LiBr) specifically in ELISA and immunohistochemical staining, while unable to bind other irrelevant cells or antigens, such as human's squamous carcinoma cells, keratinocytes,...