Detection of brain-derived neurotrophic factor (BDNF) in rat blood and brain preparations using ELISA: Pitfalls and solutions
Detection of brain-derived neurotrophic factor (BDNF) in rat blood and brain preparations using ELISA: Pitfalls and solutions
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DOI：
10.1016/j.jneumeth.2009.12.017
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发表时间：
2010-03-15
影响因子：
3
通讯作者：
Wegener, Gregers
中科院分区：
文献类型：
--
作者：
Elfving, Betina;Plougmann, Pia Hogh;Wegener, Gregers
内源性脑源性神经营养因子（BDNF）的定量可以用酶联免疫吸附测定（ELISA）进行。虽然在许多研究中用ELISA试剂盒测定了血液和脑制剂中的BDNF，但关于血液中特别是脑中BDNF浓度测量的方法学问题仅进行了表面研究。我们的目的是验证和优化BDNF ELISA试剂盒在大鼠血液和大脑样本中的测量。我们发现预分析条件对于血浆样本至关重要，但对于血清或全血样本则不然。采用最佳稀释度的额叶皮层和海马提取液，对BDNF ELISA试剂盒在大鼠血清和脑组织中的批内和批间变异性、准确度和产率进行了研究。额叶皮层和海马提取物的最佳稀释度被确定为20和120倍，并且我们确定了海马的批内变异系数（CV%）为8，额叶皮层和血清为2。海马、额叶皮质和血清中的试验间差异也很低，CV%为11或更低。最后，我们发现BDNF测量的准确性和产量在血清中较高，而在海马和额叶皮质中较低。结论：BDNF ELISA试剂盒测定血清BDNF准确，重复性好。此外，当采取特别的预防措施时，特别是注意脑组织样品的稀释时，它可用于测量大鼠脑制备物中的BDNF。(C)2009 Elsevier B.V.保留所有权利。
Quantification of endogenous brain-derived neurotrophic factor (BDNF) can be performed with an enzyme-linked immunosorbent assay (ELISA). Although BDNF has been determined in blood and brain preparations in numerous studies with ELISA kits, the methodological issues regarding measurements of BDNF concentrations in the blood and particularly in the brain have only been superficially investigated. We aimed at validating and optimizing the BDNF ELISA kit with respect to measurements in rat blood and brain samples. We found that the pre-analytical conditions were critical for plasma samples, but not serum or whole blood samples. The intra- and inter-assay variation and the accuracy and yield of the BDNF ELISA kit in rat serum and brain tissue were conducted with the optimal dilutions of frontal cortex and hippocampus extract. The optimal dilutions of frontal cortex and hippocampus extracts were determined to be 20 and 120 times, and we established that the intra-assay coefficient of variation (CV%) was 8 in hippocampus and 2 in frontal cortex and serum. The inter-assay variation was also low with a CV% of 11 or less in hippocampus, frontal cortex, and serum. Finally, we found that the accuracy and yield of the BDNF measurements were high in serum and low in hippocampus and frontal cortex. We conclude that the BDNF ELISA kit determines serum BDNF accurately and with high reproducibility. Furthermore it can be used for measurement of BDNF in rat brain preparations when particular precautions are taken and in particular with care regarding the dilution of the brain tissue samples. (C) 2009 Elsevier B.V. All rights reserved.