噬菌体抗体库技术构建HPV 16 E7人源化单克隆抗体

Construction of human monoclonal antibody to human papillomavirus type 16 E7 by phage display technology

目的：构建抗ＨＰＶ１６Ｅ７的噬菌体抗体库。方法：由感染者抗凝血中分离外周淋巴细胞，提取细胞总ＲＮＡ，以逆转录聚合酶链反应（ＲＴＰＣＲ），用人ＩｇＧ Ｆａｂ基因特异引物，从合成的ｃＤＮＡ中分别扩增出抗体轻链和重链可变区基因，回收纯化ＰＣＲ产物。用ＳｐｅＩ＋ＸｈｏＩ酶切重链可变区基因，ＸｂａＩ＋ＳａｃＩ酶切轻链可变区基因，先后克隆入噬菌体载体ｐＣｏｍｂ３。结果：ＰＣＲ产物经电泳证实分子量大小正确，酶切电泳证实质粒构建正确。结论：成功构建抗体轻链和重链基因克隆，建立抗ＨＰＶ１６Ｅ７噬菌体抗体库。

Objective:To construct to human monoclonal antibo dy to human papillomavirus type 16 E7 by Phage Display Technology.Method s:Total RNA was prepared from the peripheral blood lymphocytes of patien ts with HPV 16 infection by RT PCR. The human IgG Fab gene of heavy and light chains were amplified with specific primers. The combinatorial phage antib ody library was prepared by inserting both heavy and light chain Fab gene into p hagemid vector pComb3 and followed by the help of helper phage infection.R esults:The gel electrophoresis showed that the human IgG Fab gene of heav y and light chains were amplified successfully and the result of restriction enz yme identification was right.Conclusion:The results provided potenti al promise for future use of phage display library in generation of human monocl onal antibody to HPV16 E7.