Zondervan P E, De Jong A, Sorber C W, Kok L P, De Bruijn W C, Van der Kwast T H
Department of Pathology, Erasmus University, Rotterdam, Netherlands.
Histochem J. 1988 Jun-Jul;20(6-7):359-64. doi: 10.1007/BF01002730.
Microwave irradiation has been applied to reduce the immunogold staining time of ultrathin sections of Lowicryl embedded specimens. Labelling has been stimulated by microwave irradiation during incubation with 10 nm colloidal gold particles coated with either goat anti-mouse antibodies (GaM-gold) or goat anti-rabbit antibodies (GaR-gold) and has been compared with control incubations. Quantification has been performed on cytoplasmic membranes or lysosomes labelled with a primary antibody. Counting the gold particles over specific and non-specific sites in electron micrographs and electron microscopic images by IBAS 2000 revealed that irradiation of 25 microliters droplets both at 80 W and 150 W resulted in an accelerated immunogold labelling, while the non-specific background levels were not increased. A plateau level in immunogold labelling intensity was reached after 25 min incubation under microwave irradiation at 150 W as compared to 120 min incubation without microwaves. No improvement in localization sharpness of immunogold labelling on membranes was achieved by microwave irradiation. The microwave-mediated acceleration of immunogold staining may be considered as an example of a staining method with a restricted thermal action on microvolumes as indicated by direct temperature measurements using a fibre-optic thermometer.
微波辐射已被用于缩短用Lowicryl包埋标本的超薄切片的免疫金染色时间。在用包被有山羊抗小鼠抗体（GaM-金）或山羊抗兔抗体（GaR-金）的10纳米胶体金颗粒孵育期间，微波辐射刺激了标记，并与对照孵育进行了比较。对用一抗标记的细胞质膜或溶酶体进行了定量分析。通过IBAS 2000对电子显微镜照片和电子显微镜图像中的特异性和非特异性位点上的金颗粒进行计数，结果显示，在80瓦和150瓦功率下对25微升液滴进行辐射均导致免疫金标记加速，而非特异性背景水平并未增加。与无微波孵育120分钟相比，在150瓦微波辐射下孵育25分钟后达到了免疫金标记强度的平稳水平。微波辐射并未提高膜上免疫金标记的定位清晰度。如使用光纤温度计直接测量温度所示，微波介导的免疫金染色加速可被视为一种对微体积具有有限热作用的染色方法的实例。
