Near-Infrared Squaraine Dyes For Fluorescence Imaging In Living Cells

Near infrared dyes can effectively avoid the spontaneous absorption and fluorescence of biological samples. Squaraine is a family of functional organic dyes with absorption and fluorescence emission in near infrared region. Therefore, their desirable optical properties make them especially suitable in design fluorescent probes and used for bioimaging. Base on previous work by our group, we investigated the application of adamantyl modified squaraine for intracellular pH detection. Furthermore, two monothiosquaraine were designed, synthesized and used for intracellular Hg2+monitoring.(1) pH plays an important role in maintaining cell normal function. Acid environment in cell is usually associated with disease or dysfunction. Thus, the design intracellular pH fluorescent probe has attracted much attention. Adamantyl modified squaraine SQ3 has poor solubility in aqueous solution, so β-CD was added to improve the solubility of SQ3 as molecule container. Due to the strong interaction, β-CD and SQ3 form a supramolecular complex SQ3 (?) β-CD. Both ESI-MS and Benesi-Hildebrand analysis showed that SQ3 form 1:2 stoichiometric complexes with β-CD, and the binding constant was calculated to be 4.7×106 M-2. SQ3(?)β-CD has been characterized as fluorescent probe for pH with pKa of 6.3. The alternative addition of NaOH and HC1 into SQ3 (?) β-CD aqueous solution demonstrated that this pH probe has a good reversibility. More importantly, confocal fluorescence microscopic imaging technique allows SQ3(?)cβ-CD to detect changes of intracellular pH in living cells. Acid environment in cell led a significant fluorescence emission.(2) Due to high affinity between mercury and thiol-containing biomolecules, Hg2+ has serious biological toxicity. A squaraine MTSQ-1 containing one thiocarbonyl was designed and synthesized as a chemodosimeter for Hg2+. Due to the poor solubility, MTSQ-1 can spontaneously aggregate in aqueous media and cause substantial fluorescence quenching. So β-CD was serves as molecule container to improve the solubility of SQ3 and form a supramolecular complex MTSQ-1 β-CD. Both ESI-MS and Benesi-Hildebrand analysis showed that MTSQ-2 form a 1:2 stoichiometric complex with β-CD, and the binding constant was calculated to be 5.0×106 M-2. Hg2+, a well-known thiophile, can induce desulfurization of thiocarbonyl group. Thus, MTSQ-1 cz(?)β-CD was used to detect Hg2+ in pH 7.0 PB buffer. The detection limit was calculated to be 3.6×10-9 M (3σ/k). Confocal fluorescence microscopic imaging indicated that SQ3 c P-CD was suitable for intracellular Hg2+ detection. The cells treated with Hg2+ have significant fluorescence emission.(3) A novel water-soluble monothiosquarane MTSQ-2 linked with 2-(2-methoxyethoy)ethoxy was synthesized. In pH 7.0 PBS buffer solution, MTSQ-2 spontaneously assembled to form aggregates and caused significant fluorescence quenching. Due to Hg2+-induced desulfurization, addition of Hg2+ into MTSQ-2 solution resulted fluorescence significant enhanced. The detection limit of MTSQ-2 in PBS solution was calculated to be 1.2×10-9 M (3σ/k). The mechanism studies clearly indicated that MTSQ-2 reacted with Hg2+ to generate SQ5. The water-solubility of SQ5 is better than MTSQ-2, the fluorescence was released by disaggregation of the oxygen product. The probe was further used for intracellular Hg2+ detection.