You Mu, Liu Wen, Xu Zhen-shan, Song Li-hua
Department of Medicine, Huainan Union University, Huainan, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2012 Nov;28(11):1177-81.
To prepare and characterize monoclonal antibodies against matrix metalloproteinase-2 (MMP-2), check its expression in the tissues of human ovarian cancer and transplanted tumors in nude mice.
MMP-2 were linked to the carrier protein bovine serumalbumin (BSA) and keyhole limpet hemocyanin (KLH) using glutaraldehyde method to obtain MMP-2-BSA and MMP-2-KLH, respectively. The anti-MMP-2 monoclonal antibody was obtained through hybridoma technique. We established the cell strains secreting mAb by hybridoma technique and prepared the mAb by induction of ascites in vivo. The prepared mAb was purified by salting out with ammonium sulfate and identified by ELISA and Western blotting. We compared the mAb and commercial polyclonal antibody by immunohistochemistry and detected the expressions of MMP-2 and CA125 in ovarian cancer issues and transplanted tumor.
The artificial antigen and 3 hybridoma cell lines secreting monoclonal antibodies (mAb) against MMP-2 were obtained. The subclasses of mAb were all IgG1. The titer of peritoneal exudates was 1:1×10(6);. The expressions of MMP-2 and CA125 in transplanted tumor and ovarian cancer tissues were all high. The positive expression rate of MMP-2 checked using generated antibody was 71.2%(57/80) in ovarian cancer tissues and 16.67% (5/30) in normal tissues, with significant difference between them (P<0.01). In early stage, the positive rate of MMP-2 and CA125 combined detection was higher than that of CA125 detection alone (P<0.01). The mAb was suitable for detecting the expression of MMP-2 in human tissues and gave results consistent with commercial polyclonal antibody. The mAb was more specific than commercial mAb (P<0.01).
The anti-human MMP-2 mAb is successfully prepared, which may serve as a valuable tool in the functionaI studies of ovarian cancer.
制备抗基质金属蛋白酶-2（MMP-2）单克隆抗体并进行鉴定，检测其在人卵巢癌组织及裸鼠移植瘤中的表达。
采用戊二醛法将MMP-2分别与载体蛋白牛血清白蛋白（BSA）和钥孔戚血蓝蛋白（KLH）连接，分别获得MMP-2-BSA和MMP-2-KLH。通过杂交瘤技术获得抗MMP-2单克隆抗体。利用杂交瘤技术建立分泌单克隆抗体的细胞株，并通过体内诱导腹水制备单克隆抗体。用硫酸铵盐析法纯化制备的单克隆抗体，并用ELISA和Western印迹法进行鉴定。通过免疫组化比较该单克隆抗体与市售多克隆抗体，并检测卵巢癌组织和移植瘤中MMP-2和CA125的表达。
获得了人工抗原及3株分泌抗MMP-2单克隆抗体（mAb）的杂交瘤细胞系。单克隆抗体的亚类均为IgG1。腹水效价为1:1×10(6)。移植瘤和卵巢癌组织中MMP-2和CA125的表达均较高。用所制备抗体检测MMP-2在卵巢癌组织中的阳性表达率为71.2%（57/80），在正常组织中为16.67%（5/30），两者差异有统计学意义（P<0.01）。早期MMP-2与CA125联合检测的阳性率高于单独检测CA125（P<0.01）。该单克隆抗体适用于检测人组织中MMP-2的表达，结果与市售多克隆抗体一致。该单克隆抗体比市售单克隆抗体更具特异性（P<0.01）。
成功制备了抗人MMP-2单克隆抗体，可为卵巢癌的功能研究提供有价值的工具。
