一种用于汗液吡啶啉和脱氧吡啶啉交联物的收集方法及高灵敏度酶免疫测定法。

A collection method and high-sensitivity enzyme immunoassay for sweat pyridinoline and deoxypyridinoline cross-links.

作者信息

Sarno M, Powell H, Tjersland G, Schoendorfer D, Harris H, Adams K, Ogata P, Warnick G R

机构信息

Vision Biotechnology Consulting, 306-N W El Norte Pkwy., PMB 311, Escondido, CA 92026, USA.

出版信息

Clin Chem. 1999 Sep;45(9):1501-9.

PMID:10471653

Abstract

BACKGROUND

Collagen cross-link molecules such as pyridinoline (PYD), deoxypyridinoline (DPD), and N-terminal cross-linked peptides (NTX) have been measured in urine as indices of bone resorption. However, very little is known regarding the excretion of pyridinolines into other biological fluids. We report a collection device, normalizing analyte, and high-sensitivity immunoassay for quantitative analysis of free pyridinoline cross-links in sweat.

METHODS

Flame atomic emission and ion-selective electrode techniques were used to measure potassium as a sweat volume marker. The Pyrilinks immunoassay for urine free pyridinolines was optimized to increase sensitivity for measurements in sweat. The precision, accuracy, and detection limit of this assay were characterized. To assess values and variability of sweat pyridinolines in human subjects, a nonocclusive skin patch was used to collect sweat samples from a reference group and from a mixed group experiencing accelerated bone resorption, postmenopausal women and men receiving gonadotropin-releasing hormone for prostate cancer.

RESULTS

The immunoassay intra- and interassay variations were </=10% and <16%, respectively, with a detection limit of 309 pmol/L. Linearity upon dilution and analytical recovery ranged from 93% to 109% and 85% to 122%, respectively. Sweat PYD values normalized to potassium output yielded a weekly intraindividual biological variability of 14.7%. The mean increase in the population experiencing increased bone resorption vs the reference group was 36% (P <0.05) for sweat PYD/K vs 23-40% (P <0.05) for urinary PYD/Cr, DPD/Cr, and NTX/Cr.

CONCLUSION

We conclude that this new platform sweat collection technology and PYD immunoassay show potential as an indicator of bone resorption.

摘要

背景

诸如吡啶啉（PYD）、脱氧吡啶啉（DPD）和N端交联肽（NTX）等胶原交联分子已在尿液中进行检测，作为骨吸收的指标。然而，关于吡啶啉在其他生物体液中的排泄情况知之甚少。我们报告了一种用于定量分析汗液中游离吡啶啉交联物的收集装置、标准化分析物和高灵敏度免疫测定法。

方法

采用火焰原子发射和离子选择性电极技术测定钾作为汗液体积标志物。对用于检测尿游离吡啶啉的Pyrilinks免疫测定法进行优化，以提高在汗液中测量的灵敏度。对该测定法的精密度、准确性和检测限进行了表征。为了评估人体汗液中吡啶啉的数值和变异性，使用非封闭性皮肤贴片从一个参考组以及一组经历骨吸收加速的混合组（绝经后妇女和接受促性腺激素释放激素治疗前列腺癌的男性）收集汗液样本。

结果

免疫测定法的批内和批间变异分别≤10%和<16%，检测限为309 pmol/L。稀释后的线性度和分析回收率分别为93%至109%和85%至122%。以钾排出量标准化的汗液PYD值产生的个体内每周生物学变异为14.7%。骨吸收增加人群与参考组相比，汗液PYD/钾的平均增加为36%（P<0.05），而尿PYD/肌酐、DPD/肌酐和NTX/肌酐的平均增加为23%至40%（P<0.05）。