Visa N, Fibla J, Santa-Cruz M C, Gonzàlez-Duarte R
Departament de Genètica, Facultat de Biologia, Universitat de Barcelona, Spain.
J Histochem Cytochem. 1992 Jan;40(1):39-49. doi: 10.1177/40.1.1729353.
To analyze Drosophila alcohol dehydrogenase gene (Adh) expression and tissue distribution at various developmental stages, we devised several immunochemical techniques making use of monoclonal antibodies against Drosophila alcohol dehydrogenase (ADH), which had been obtained previously. We here report their application to analyze the expression of Adh in a wild-type strain of D. melanogaster. s-ELISA tests were performed to evaluate fluctuations in ADH content and specific activity during development in individual organs as well as in whole individuals. In all cases, ADH specific activity appeared to be quite constant, which implies that variations in enzyme activity reflect differences in protein content. Immunoblottings of crude homogenates revealed immunoreactive low relative molecular mass peptides in addition to the 27 KD monomeric band, showing a conserved banding pattern in different organs and developmental stages. Immunohistochemical assays on whole organs were used to analyze the general pattern of ADH distribution. Immunoperoxidase staining of cryosections proved to be of crucial relevance, as it yielded full details of the tissue localization of ADH within the ADH-positive organs. We have shown not only that ADH displays a specific distribution in some organs but also that the enzyme is restricted to certain cell types.
为了分析果蝇乙醇脱氢酶基因（Adh）在不同发育阶段的表达及组织分布，我们利用先前获得的抗果蝇乙醇脱氢酶（ADH）的单克隆抗体设计了几种免疫化学技术。我们在此报告它们在分析黑腹果蝇野生型品系中Adh表达方面的应用。进行了s-ELISA试验，以评估个体器官以及整个个体发育过程中ADH含量和比活性的波动。在所有情况下，ADH比活性似乎相当恒定，这意味着酶活性的变化反映了蛋白质含量的差异。粗匀浆的免疫印迹显示，除了27 KD的单体条带外，还有免疫反应性低相对分子质量肽段，在不同器官和发育阶段呈现出保守的条带模式。对整个器官进行免疫组织化学分析以研究ADH分布的总体模式。冷冻切片的免疫过氧化物酶染色被证明至关重要，因为它提供了ADH在ADH阳性器官内组织定位的完整细节。我们不仅表明ADH在某些器官中呈现特定分布，还表明该酶局限于某些细胞类型。
