Monoclonal antibody assay for free urinary pyridinium cross-links.
Monoclonal antibody assay for free urinary pyridinium cross-links.
复制标题
用于游离尿吡啶交联的单克隆抗体测定。
作者：
B. Gomez;Jr.;S. Ardakani;B. Evans;Loren D. Merrell;D. Jenkins;V. T. Kung
胶原蛋白的吡啶交联，吡啶啉（Pyd）和脱氧吡啶啉（Dpd），提供了骨胶原原纤维的结构完整性和刚性。在骨降解（再吸收）过程中，交联被释放到血液循环中，最终通过尿液排出体外。尿中吡啶交联测量已被证明是敏感和特异性的指标吸收建立高效液相色谱和较新的酶免疫测定（EIA）技术。我们已经开发出一种单克隆抗体，可以优先结合非肽结合的Pyd和Dpd的自由形式。我们利用pyd包被微滴带孔将抗体偶联到碱性磷酸酶的竞争性环评中。样品和抗体-酶偶联物孵育3小时后，以对硝基苯磷酸为底物，1小时显色。试验内（n = 52） cv为3.0 ~ 7.6%，试验间（n = 8） cv为6.1 ~ 7.4%。将该方法(y)与HPLC (x)和基于多克隆抗体的EIA （x’）进行比较，得到的回归方程为y = 0.46x + 4, r = 0.96; y = 0.56x’+ 8,r = 0.96。EIA检测到绝经后妇女和骨质疏松症、甲亢、甲状旁腺功能亢进和骨佩吉特病患者尿液中Pyd和Dpd浓度升高。EIA浓度也反映了手术绝经妇女雌激素替代导致的pyd和dpd排泄减少。用这种简单的EIA测量吡啶交联似乎提供了一个准确的骨吸收率指数，可能对代谢性骨病的评估和监测抗骨吸收治疗的效果有用。
The pyridinium cross-links of collagen, pyridinoline (Pyd) and deoxypyridinoline (Dpd), provide structural integrity and rigidity to collagen fibrils in bone. During bone degradation (resorption), the cross-links are released into the circulation and eventually excreted in urine. Pyridinium cross-link measurements in urine have been shown to be sensitive and specific indicators of resorption by both established HPLC and newer enzyme immunoassay (EIA) techniques. We have developed a monoclonal antibody that preferentially binds to the non-peptide-bound free forms of Pyd & Dpd. We have incorporated the antibody conjugated to alkaline phosphatase in a competitive EIA by using Pyd-coated microtiter strip wells. After a 3-h incubation of sample and antibody-enzyme conjugate, color is developed for 1 h with p-nitrophenyl phosphate as the substrate. The intraassay (n = 52) CVs were 3.0-7.6%, and interassay (n = 8) CVs were 6.1-7.4%. Comparisons of the assay (y) with HPLC (x) and a polyclonal antibody-based EIA (x') gave regression equations of y = 0.46x + 4, r = 0.96, and y = 0.56x' + 8, r = 0.96. The EIA detected increased Pyd & Dpd concentrations in urine from postmenopausal women and patients with osteoporosis, hyperthyroidism, hyperparathyroidism, and Paget disease of bone. EIA concentrations also reflected the reduction in Pyd&Dpd excretion resulting from estrogen replacement in surgically menopausal women. Measurement of pyridinium cross-links with this simple EIA appears to provide an accurate index of the rate of resorption and may be useful for metabolic bone disease assessment and monitoring the effects of antiresorptive therapy.
影响因子：
4.1
作者：
EYRE, DR;DICKSON, IR;VANNESS, K
通讯作者：
VANNESS, K