Centre for Psychiatric Research, Aarhus University Hospital, Skovagervej 2, DK-8240 Risskov, Denmark.
J Neurosci Methods. 2010 Mar 15;187(1):73-7. doi: 10.1016/j.jneumeth.2009.12.017. Epub 2010 Jan 5.
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.
内源性脑源性神经营养因子（BDNF）的定量可以通过酶联免疫吸附测定（ELISA）进行。尽管已有许多研究使用 ELISA 试剂盒在血液和脑组织中测定了 BDNF，但在血液中，尤其是在大脑中测量 BDNF 浓度的方法学问题仅得到了初步研究。我们旨在验证和优化 BDNF ELISA 试剂盒，以用于大鼠血液和脑组织样本的测量。我们发现，血浆样品的分析前条件至关重要，但血清或全血样品则不然。使用最佳稀释度的前额皮质和海马提取物进行了大鼠血清和脑组织中内、间分析变异以及 BDNF ELISA 试剂盒的准确性和产率的检测。确定最佳稀释度的前额皮质和海马提取物分别为 20 倍和 120 倍，我们还确定海马内的内分析变异系数（CV%）为 8，前额皮质和血清中的 CV%为 2。海马、前额皮质和血清中的间分析变异性也较低，CV%低于 11。最后，我们发现血清中的 BDNF 测量准确性和重复性高，而海马和前额皮质中的 BDNF 测量准确性和重复性较低。我们的结论是，BDNF ELISA 试剂盒能够准确、高度重现地测定血清中的 BDNF。此外，当采取特殊预防措施且特别注意脑组织样品的稀释度时，它可以用于测量大鼠脑组织中的 BDNF。
