Jakab Balázs, Reglodi Dóra, Józsa Rita, Hollósy Tibor, Tamás Andrea, Lubics Andrea, Lengvári István, Oroszi Gábor, Szilvássy Zoltán, Szolcsányi János, Németh József
Neuropharmacology Research Group of the Hungarian Academy of Sciences, Department of Pharmacology and Pharmacotherapy, University of Pécs, Szigeti út 12, H-7624 Pécs, Hungary.
J Biochem Biophys Methods. 2004 Oct 29;61(1-2):189-98. doi: 10.1016/j.jbbm.2004.03.002.
Pituitary adenylate cyclase activating polypeptide (PACAP) occurs in two molecular forms: PACAP-38 and PACAP-27. Soon after the isolation and chemical characterization of PACAP, the first radioimmunoassay (RIA) methods have been developed, but it is a still rarely used laboratory technique in the field of PACAP research. The aim of the present study was to develop a novel, highly specific PACAP-38 assay to investigate the quantitative distribution of PACAP-38 in the central nervous system of various vertebrate species under the same technical and experimental conditions. Different areas of the brain and the spinal cord were removed from rats, chickens and fishes and the tissue samples were processed for PACAP-38 RIA. Our results indicate that the antiserum used in the RIA is C-terminal specific, without affinity for other members of the vasoactive intestinal polypeptide (VIP)/secretin/glucagon peptide family. The average ID50 value was 48.6+/-3.4 fmol/ml determined in 10 consecutive assays. Detection limit for PACAP-38 proved to be 2 fmol/ml. PACAP-38 immunoreactivity was present in the examined brain areas of each species studied, with highest concentration in the rat diencephalons. High levels of PACAP-38 were also detected in the rat telencephalon, followed by spinal cord and brainstem. The central nervous system of the fish also contained considerable concentrations of PACAP-38, whereas lowest concentrations were measured in the central nervous system of the chicken.
垂体腺苷酸环化酶激活多肽（PACAP）以两种分子形式存在：PACAP - 38和PACAP - 27。在PACAP被分离并进行化学表征后不久，就开发出了首批放射免疫分析（RIA）方法，但在PACAP研究领域，这仍是一种很少使用的实验室技术。本研究的目的是开发一种新型、高特异性的PACAP - 38检测方法，以在相同的技术和实验条件下研究PACAP - 38在各种脊椎动物物种中枢神经系统中的定量分布。从大鼠、鸡和鱼身上取出大脑和脊髓的不同区域，对组织样本进行PACAP - 38 RIA处理。我们的结果表明，RIA中使用的抗血清是C端特异性的，对血管活性肠肽（VIP）/促胰液素/胰高血糖素肽家族的其他成员没有亲和力。在连续10次测定中，平均ID50值为48.6±3.4 fmol/ml。PACAP - 38的检测限被证明为2 fmol/ml。在所研究的每个物种的检测脑区中都存在PACAP - 38免疫反应性，在大鼠间脑中浓度最高。在大鼠端脑中也检测到高水平的PACAP - 38，其次是脊髓和脑干。鱼的中枢神经系统中也含有相当浓度的PACAP - 38，而在鸡的中枢神经系统中测得的浓度最低。
