Discipline of Medicine, Central Clinical School.
Discipline of Child and Adolescent Health, Children's Hospital at Westmead Clinical School, Faculty of Medicine and Health, The University of Sydney, Sydney, NSW, Australia.
Appl Immunohistochem Mol Morphol. 2021 Jul 1;29(6):e46-e56. doi: 10.1097/PAI.0000000000000925.
Neuronal nuclei (NeuN) is a neuron-specific nuclear protein, reported to be stably expressed in most postmitotic neurons of the vertebrate nervous system. Reduced staining has been interpreted by some to indicate loss of cell viability in human studies, while others suggest this may be because of changes in the antigenicity of the target epitope. Preliminary studies in our laboratory found low immunostaining for the NeuN antibody on formalin fixed and paraffin embedded (FFPE) human brain tissue. We report on the techniques and results used to enhance the staining for NeuN in that tissue. In parallel, we stained NeuN in piglet brain tissue, sourced from an experimental model where methodological parameters, including those for tissue fixation and storage, were tightly controlled. In human FFPE brain tissue, we were unable to enhance NeuN immunostaining to a degree sufficient for cell counting. In contrast, we found consistently high levels of staining in the piglet brain tissue. We conclude that processes used for fixation and storage of human FFPE brain tissue are responsible for the reduced staining. These results emphasize that a cautionary approach should be taken when interpreting NeuN staining outcomes in human FFPE brain tissue.
神经元核（NeuN）是一种神经元特异性核蛋白，据报道在脊椎动物神经系统的大多数有丝分裂后神经元中稳定表达。一些人认为染色减少表明人类研究中细胞活力丧失，而另一些人则认为这可能是因为靶抗原表位的抗原性发生变化。我们实验室的初步研究发现，福尔马林固定和石蜡包埋（FFPE）的人脑组织中 NeuN 抗体的免疫染色较弱。我们报告了用于增强该组织中 NeuN 染色的技术和结果。同时，我们对来自实验模型的仔猪脑组织中的 NeuN 进行了染色，该模型中严格控制了包括组织固定和储存在内的方法学参数。在人类 FFPE 脑组织中，我们无法增强 NeuN 免疫染色的程度，不足以进行细胞计数。相比之下，我们发现仔猪脑组织中的染色始终很高。我们得出结论，用于固定和储存人类 FFPE 脑组织的过程是导致染色减少的原因。这些结果强调，在解释人类 FFPE 脑组织中的 NeuN 染色结果时应采取谨慎的方法。
