权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Immunohistochemical and immunocytochemical studies of gibberellins

赤霉素的免疫组织化学和免疫细胞化学研究

基本信息

批准号：
03453142
负责人：
MUROFUSHI Noboru
金额：
$ 4.99万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1992
项目状态：
已结题

项目摘要

For the immunohistochemistry and immunocytochemistry of gibberellins and GA-binding proteins, monoclonal antibody against GA19 and GA24, and anti-idiotypic antibody against a monoclonal anti-GA4-antibody were prepared. To determine the procedure for fixation of gibberellins, treatments of rapid freezing-freeze substitution in osmate/acetone/dry-ice, and of freeze-drying followed by fixation with gaseous osmate were examined using rice anthers as a plant material, and clear fixation of organelle was observed in the rapid freezing-freeze substitution procedure. The well fixed rice anthers were stained with anti-gibberellin- antibody as the first antibody followed by colloidal gold labeled rabbit anti-mouse IgG. When anti-GA19/24-antibody was used as the first antibody, mitochondria, nuclei (chromatin in particular) and cuticle in epidermal cells of anther, mitochondria, chloroplasts, nuclei (chromatin in particular) and orbiculus in tapetal cells of anther, exine of cell wall, mitochondria and polysaccharide particles in pollen were stained. They were not stained with normal mouse IgG as the first antibody. When anti-GA19/24-antibody preincubated with GA19 was used for immunostaining, nothing was stained, suggesting the specific immunostaining of the above tissues and organelles with anti-GA19/24-antibody. When the anther was stained with anti- GA4-antibody as the first antibody, similar tissues and organelles were stained, but the staining was still observed when stained with the antibody preincubated with GA4 or GA1.

为了进行赤霉素和GA结合蛋白的免疫组织化学和免疫细胞化学，制备了针对GA 19和GA 24的单克隆抗体以及针对单克隆抗GA 4抗体的抗独特型抗体。以水稻花药为材料，研究了渗透压/丙酮/干冰快速冷冻-冷冻置换法和冷冻干燥后渗透压气体固定法对赤霉素固定的影响。以抗赤霉素抗体为第一抗体，胶体金标记兔抗鼠IgG，对固定好的水稻花药进行染色。当用抗GA 19/24抗体作为第一抗体时，花药表皮细胞中的线粒体、细胞核（特别是染色质）和角质层，花药绒毡层细胞中的线粒体、叶绿体、细胞核（特别是染色质）和圆形体，细胞壁外壁，花粉中的线粒体和多糖颗粒被染色。它们不被作为第一抗体的正常小鼠IgG染色。当使用与GA 19预孵育的抗GA 19/24-抗体进行免疫染色时，没有染色，表明抗GA 19/24-抗体对上述组织和细胞器进行特异性免疫染色。当花药用抗GA 4抗体作为第一抗体染色时，类似的组织和细胞器被染色，但当用GA 4或GA 1预孵育的抗体染色时仍观察到染色。