‡Laboratory of Bio-imaging Probe Development, Singapore Bio-imaging Consortium, Agency of Science, Technology and Research, Singapore 138667, Singapore.
J Am Chem Soc. 2015 Apr 29;137(16):5355-62. doi: 10.1021/ja5115776. Epub 2015 Apr 21.
Two-photon (TP) microscopy has an advantage for live tissue imaging which allows a deeper tissue penetration up to 1 mm comparing to one-photon (OP) microscopy. While there are several OP fluorescence probes in use for pancreatic islet imaging, TP imaging of selective cells in live islet still remains a challenge. Herein, we report the discovery of first TP live pancreatic islet imaging probe; TP-α (Two Photon-alpha) which can selectively stain glucagon secreting alpha cells. Through fluorescent image based screening using three pancreatic cell lines, we discovered TP-α from a TP fluorescent dye library TPG (TP-Green). In vitro fluorescence test showed that TP-α have direct interaction and appear glucagon with a significant fluorescence increase, but not with insulin or other hormones/analytes. Finally, TP-α was successfully applied for 3D imaging of live islets by staining alpha cell directly. The newly developed TP-α can be a practical tool to evaluate and identify live alpha cells in terms of localization, distribution and availability in the intact islets.
双光子（TP）显微镜在活体组织成像方面具有优势，与单光子（OP）显微镜相比，其允许更深的组织穿透深度可达 1 毫米。虽然有几种用于胰腺胰岛成像的 OP 荧光探针，但活体胰岛中选择性细胞的 TP 成像仍然是一个挑战。在这里，我们报告了首例用于活体胰腺胰岛成像的 TP 探针的发现；TP-α（双光子-α），它可以选择性地染色胰高血糖素分泌的α细胞。通过使用三种胰腺细胞系进行基于荧光图像的筛选，我们从 TP 荧光染料库 TPG（TP-Green）中发现了 TP-α。体外荧光试验表明，TP-α 具有直接相互作用，并与胰高血糖素出现明显的荧光增强，但与胰岛素或其他激素/分析物没有。最后，TP-α 成功地应用于通过直接染色α细胞对活体胰岛进行 3D 成像。新开发的 TP-α 可以成为一种实用的工具，用于评估和识别完整胰岛中定位、分布和可用性的活α细胞。
