Evaluation of the specificity of antibodies raised against cannabinoid receptor type 2 in the mouse retina
Evaluation of the specificity of antibodies raised against cannabinoid receptor type 2 in the mouse retina
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DOI：
10.1007/s00210-013-0930-8
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发表时间：
2014-02-01
影响因子：
3.6
通讯作者：
Bouchard, Jean-Francois
中科院分区：
文献类型：
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作者：
Cecyre, Bruno;Thomas, Sebastien;Bouchard, Jean-Francois
大麻素受体（CB 1 R和CB 2 R）是中枢神经系统中最丰富的G蛋白偶联受体之一。内源性大麻素系统是免疫系统调节和外周疼痛管理的有吸引力的治疗靶点。虽然CB 1 R分布在神经系统中，但CB 2 R传统上与免疫系统相关。自从使用基于抗体的方法在神经元中发现CB 2 R表达以来，这一教条目前是一个争论的主题。CB 2 R在中枢神经系统（CNS）中的定位可能对药物开发产生重大影响，因为这意味着除了对外周疼痛通路的影响外，CB 2 R还可以介导大麻素的一些中枢作用。为了澄清关于CB 2 R在CNS中表达的争论，我们使用Western印迹和免疫组织化学对从野生型小鼠和缺乏CB 2 R（cnr 2（-/-））的小鼠获得的视网膜组织测试了几种商业上或学术上生产的CB 2 R抗体。测试的抗体之一表现出有价值的特异性，因为它在Western印迹中标记接近预测分子量的单个条带，并且在cnr 2（-/-）小鼠视网膜切片中不产生染色。当与它们的免疫肽一起使用或在cnr 2（-/-）视网膜切片上使用时，测试的其他抗体在Western印迹中检测到多个条带并标记未鉴定的蛋白质。我们的结论是，许多常用的抗CB 2 R的抗体在免疫组织化学中的使用是不特异的，至少在小鼠视网膜的情况下。此外，其中一些受试品存在显著的批间变异性。因此，在解释使用CB 2 R抗体的既往和未来研究时应谨慎。
Cannabinoid receptors (CB1R and CB2R) are among the most abundant G protein-coupled receptors in the central nervous system. The endocannabinoid system is an attractive therapeutic target for immune system modulation and peripheral pain management. While CB1R is distributed in the nervous system, CB2R has traditionally been associated to the immune system. This dogma is currently a subject of debate since the discovery of CB2R expression in neurons using antibody-based methods. The localization of CB2R in the central nervous system (CNS) could have a significant impact on drug development because it would mean that in addition to its effects on the peripheral pain pathway, CB2R could also mediate some central effects of cannabinoids. In an attempt to clarify the debate over CB2R expression in the CNS, we tested several commercially or academically produced CB2R antibodies using Western blot and immunohistochemistry on retinal tissue obtained from wild-type mice and mice lacking CB2R (cnr2 (-/-) ). One of the antibodies tested exhibited a valuable specificity as it marked a single band near the predicted molecular weight in Western blot and produced no staining in cnr2 (-/-) mice retina sections. The other antibodies tested detected multiple bands in Western blot and labeled unidentified proteins when used with their immunizing peptide or on cnr2 (-/-) retinal sections. We conclude that many commonly used antibodies raised against CB2R are not specific for use in immunohistochemistry, at least in the context of the mouse retina. Moreover, some of them tested presented significant lot-to-lot variability. Hence, caution should be used when interpreting prior and future studies using CB2R antibodies.