Ebinuma Hiroyuki, Miida Takashi, Yamauchi Toshimasa, Hada Yusuke, Hara Kazuo, Kubota Naoto, Kadowaki Takashi
Diagnostics Research Laboratories, Daiichi Pure Chemicals Co. Ltd., Ibaraki, Japan.
Clin Chem. 2007 Aug;53(8):1541-4. doi: 10.1373/clinchem.2007.085654. Epub 2007 Jun 28.
Human serum adiponectin exists in 3 multimer forms: high molecular weight (HMW), middle molecular weight, and low molecular weight (LMW), with some of the latter bound to albumin (Alb)-LMW. Some studies have suggested that adiponectin crosses the blood-brain barrier and plays a central role in energy homeostasis.
To determine cerebrospinal fluid (CSF) adiponectin at extremely low concentrations, we modified the protocol of the ELISA system used to assay serum adiponectin. The 3 multimers of adiponectin were measured separately by pretreating CSF with 2 proteases. We measured the CSF adiponectin concentrations in anonymous human samples (n = 19). The molecular sizes of adiponectin in CSF pretreated with proteases or untreated were determined by use of native PAGE and immunoblotting.
The ELISA system measured adiponectin in the range of 1.0-167 microg/L. The between-assay imprecision estimates (CVs) were 6%-17% for the 3 forms. The mean total CSF adiponectin concentration (7.2 microg/L) was approximately 1/1000 of the mean concentration in serum. Unlike serum adiponectin, the LMW and Alb-LMW forms predominated in all of the CSF samples. Immunoblotting analysis revealed that most LMW forms were bound to Alb, although the HMW form was detected in some samples.
The modified ELISA system measures the 3 multimers separately and is sufficiently sensitive to measure adiponectin in CSF.
人血清脂联素以三种多聚体形式存在：高分子量（HMW）、中分子量和低分子量（LMW），其中一些低分子量形式与白蛋白（Alb）结合形成Alb-LMW。一些研究表明，脂联素可穿过血脑屏障并在能量稳态中起核心作用。
为了测定极低浓度的脑脊液（CSF）脂联素，我们修改了用于检测血清脂联素的ELISA系统方案。通过用两种蛋白酶预处理脑脊液，分别测量脂联素的三种多聚体。我们测量了匿名人类样本（n = 19）中的脑脊液脂联素浓度。使用天然PAGE和免疫印迹法测定经蛋白酶预处理或未处理的脑脊液中脂联素的分子大小。
ELISA系统测量的脂联素浓度范围为1.0 - 167μg/L。三种形式的批间不精密度估计值（CV）为6% - 17%。脑脊液中脂联素的平均总浓度（7.2μg/L）约为血清平均浓度的1/1000。与血清脂联素不同，所有脑脊液样本中低分子量和Alb-LMW形式占主导。免疫印迹分析显示，大多数低分子量形式与白蛋白结合，尽管在一些样本中检测到了高分子量形式。
改良的ELISA系统可分别测量三种多聚体，并且对测量脑脊液中的脂联素具有足够的敏感性。
