Photo-activation Localization Microscopy (PALM) is one of super-resolved fluorescence microscopies. In PALM imaging, a target is labeled with a dye exhibiting photo-switchable fluorescence. The fluorescence of the dye is randomly turned ON and OFF at a low concentration of ON-state dyes where the distance among dyes in the ON-state in a video frame is long enough to be spatially resolved by a conventional optical microscope. The detailed positions of the single fluorescence spots are determined at nanometer accuracy by localization method and, the locations of all the fluorescence spots are re-constructed to obtain a super-resolved fluorescence (PALM) image. In PALM, two or more light sources are usually used for photo-activation, excitation, and deactivation of fluorescence; this requires a relatively complicated experimental setup. In my presentation, I will show a new approach for the ON-OFF switching of fluorescence using the ‘‘hot-band’’ of a photo-switchable fluorescent diarylethene derivative (DE) and its application to super-resolved fluorescence imaging.
光の回折限界を超えた空間分解能を有する超解像顕微鏡の一つPhoto-activation Localization Microscopy (PALM)では，光によって蛍光のON-OFF制御が可能な光スイッチング蛍光分子で試料を修飾し，それらの蛍光を，単一蛍光スポット間の間隔が光学顕微鏡で空間分解できるほど広くなる条件で時空間的にランダムON-OFFさせる。孤立した単一分子の位置をnm精度で決定可能な局在化（localization）法を蛍光ON状態の色素に対して適用し，全ての蛍光分子の位置をコンピュータで再構築することで超解像イメージングを実現する。一般に，ON/OFFスイッチングにより蛍光分子数を制御するために複数光源が用いられるが，光学系が複雑になるといった課題がある。本発表では，単一波長の光照射によって蛍光の自発的ON-OFFを実現した研究成果とその詳細なメカニズム，一波長PALMへの応用を紹介する。