Rønne E, Høyer-Hansen G, Brünner N, Pedersen H, Rank F, Osborne C K, Clark G M, Danø K, Grøndahl-Hansen J
Finsen Laboratory, Rigshospitalet, Copenhagen, Denmark.
Breast Cancer Res Treat. 1995 Mar;33(3):199-207. doi: 10.1007/BF00665944.
Urokinase plasminogen activator (uPA) is a proteolytic enzyme involved in degradation of the extracellular matrix during cancer invasion. The levels of uPA and its inhibitor PAI-1 in tumor extracts have previously been demonstrated to be of prognostic value in breast cancer as well as other types of cancer. We have previously characterized a specific cell surface receptor for uPA (uPAR) which strongly enhances the catalytic activity of uPA and is expressed during mammary cancer invasion. In order to quantitate uPAR in breast cancer tissue, we have now developed a sensitive enzyme-linked immunosorbent assay (ELISA), with polyclonal catching antibodies and three monoclonal detecting antibodies. The detection limit of the assay is approximately 0.16 fmol of uPAR in a volume of 100 microliters (1.6 pM). There is a linear relationship between signal and uPAR concentration up to at least 6.6 fmol per 100 microliters (66 pM). Both free uPAR and uPAR in complex with uPA is detected. The recovery of an internal uPAR standard in breast cancer tissue extracts is above 87%. The intra-assay and inter-assay variation coefficients are 7% and 13%. In order to find a suitable buffer for extraction of various components of the uPA-system from breast cancer tissue, we tested buffers which previously have been used for optimal extraction of estrogen receptor (A), uPA (B), and uPAR (C). Buffer A and B extracted approximately 30% and 50%, respectively, of the amount of uPAR extracted with buffer C.(ABSTRACT TRUNCATED AT 250 WORDS)
尿激酶型纤溶酶原激活剂（uPA）是一种蛋白水解酶，参与癌症侵袭过程中细胞外基质的降解。肿瘤提取物中uPA及其抑制剂PAI-1的水平先前已被证明在乳腺癌以及其他类型的癌症中具有预后价值。我们之前已鉴定出一种uPA的特异性细胞表面受体（uPAR），它能强烈增强uPA的催化活性，并在乳腺癌侵袭过程中表达。为了定量乳腺癌组织中的uPAR，我们现已开发出一种灵敏的酶联免疫吸附测定法（ELISA），使用多克隆捕获抗体和三种单克隆检测抗体。该测定法的检测限在100微升体积中约为0.16飞摩尔uPAR（1.6皮摩尔）。信号与uPAR浓度之间存在线性关系，每100微升至少可达6.6飞摩尔（66皮摩尔）。游离uPAR和与uPA结合的uPAR均可被检测到。乳腺癌组织提取物中内源性uPAR标准品的回收率高于87%。批内和批间变异系数分别为7%和13%。为了找到一种适合从乳腺癌组织中提取uPA系统各成分的缓冲液，我们测试了先前用于最佳提取雌激素受体（A）、uPA（B）和uPAR（C）的缓冲液。缓冲液A和B分别提取了用缓冲液C提取的uPAR量的约30%和50%。（摘要截短于250字）
