蛋白质及其标记.pptxVIP

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蛋白质及其标记.pptx

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生物分子的化学标记和探针技术Some of the common bioconjugate designs often used for life science applications include:(A) streptavidin–enzyme conjugate(B) an immobilized affinity ligand on a particle(C) an oligo molecular beacon probe containing two fluorescent labels or a fluor and a quencher at each end(D) fluorescently labeled streptavidin(E) an affinity ligand attached to a surface (F) a biotinylated enzyme(G) an antibody–enzyme conjugate(H) a fluorescently labeled antibody(I) a biotinylated antibody(J) a biotinylated oligo probe(K) an antibody–drug conjugate(L) a gadolinium chelate-modified dendrimer containing folate molecules for targeting.蛋白质和核酸的荧光标记Localization of Molecules with Fluorescence MicroscopyImage from Molecular Probes (Invitrogen): Fluorescence Applications in BiologyFluorescence Techniques Can Provide the Following Information:Simple fluorophores:- Localization of biomolecules- Assessment of binding- Calculation of diffusion coefficientsMore complex systems:- Reporting of gene activation- Detection of physiological changes- Measurement of distances and conformational changesExamples of Fluorescent MoleculesFluorescence of native proteins residues: Tryptophan: lex = 290, lem = 330-350, depending on environmentDNA and RNA generally exhibit no fluorescence.Fluorescein (Fluorescent)lex = 494,lem = 518Phenolphthalein (Non-fluorescent)Pyrenelex = 345, lem = 3787-Hydroxycoumarinlex = 385, lem = 445Coumarin 343lex = 454, lem = 495BODIPY 493lex = 500, lem = 506TAMRAlex = 555, lem = 580Texas Redlex = 595, lem = 615Cy5lex = 643, lem = 667荧光的产生 Partial energy diagram for a photoluminescent system. Spectral observables for fluorescence sensing. From left to right: Intensity, intensity ratio, anisotropy, time-domain lifetime, and phase-modulation lifetime. 蛋白质的选择性标记Common Examples of Protein BioconjugatesNewFunctionalityChemicalReactionChromophoresCatalystsSpin LabelsAffinity LabelsCrosslinkersPolymersMaterial SurfacesMRI Contrast AgentsRadiolabelsNanocrystalsEnzym