RESUMO
To expand the applicability of the biotin-(strept)avidin system, a biotin analog with reversible binding under non-denaturing conditions has been designed, and its applications in protein labeling, detection, and isolation have been evaluated.
Assuntos
Biotina/análogos & derivados , Estreptavidina/análise , Linhagem Celular Tumoral , Corantes Fluorescentes/química , Humanos , Ligação de Hidrogênio , Microscopia Confocal , Ligação Proteica , Estreptavidina/química , Estreptavidina/isolamento & purificaçãoRESUMO
New BODIPY-based pH probes have been designed with excitation and emission wavelengths suitable for fluorescence microscopy and flow cytometry. These pH probes are cell-permeable, selectively label lysosomes, and can be used for noninvasive monitoring of lysosomal pH changes during physiological and pathological processes.
Assuntos
Equilíbrio Ácido-Base , Compostos de Boro/química , Corantes Fluorescentes/química , Lisossomos/química , Lisossomos/metabolismo , Modelos Biológicos , Animais , Compostos de Boro/síntese química , Bovinos , Linhagem Celular , Citometria de Fluxo , Concentração de Íons de Hidrogênio , Microscopia Confocal , Estrutura MolecularRESUMO
The use of genetically encoded small peptide tags such as polyhistidine and tetracysteine tags has become important for protein purification and enrichment. An improved affinity purification of tetracysteine (CCXXCC) tagged proteins has been achieved using a nonfluorescent, photochemically stable bisarsenical affinity ligand SplAsH. The photochemical stability of the SplAsH-biotin, shown in compound 5, is superior to FlAsH-EDT(2) and ReAsH-EDT(2). An application of the SplAsH tag for affinity purification of tetracysteine-tagged proteins is reported.
Assuntos
Arsenicais/química , Cisteína/análogos & derivados , Cisteína/química , Proteínas/química , Proteínas/isolamento & purificação , Arsenicais/síntese química , Biotina/química , Cromatografia de Afinidade , Cinética , Ligantes , Estrutura Molecular , Processos Fotoquímicos , EstereoisomerismoRESUMO
Efficient syntheses of several members of a new class of symmetric, monocarboxylate-functionalized cyanine dyes have been developed. The synthesis is a simple two-step method, typically with greater than 60% yield and easy final product purification. The new monocarboxylate-functionalized cyanine dyes exhibit excellent water solubility and similar excitation and emission properties to those of Cy5 and Alexa Fluor 647. The application of the new dyes in cellular imaging has been demonstrated through direct conjugating of the dye with an antibody, then imaging of microtubules inside cells, visualized by near-infrared fluorescence microscopy.
