荧光标记或荧光探针是众多成像和诊断应用的理想选择。荧光多肽的应用范围包括肽与蛋白质相互作用研究、酶活性测定或新疾病模型的开发。荧光团可以共价连接到N端或C端，但建议与N端连接。由受体和猝灭剂组成的FRET对可以在内部或外部连接。对于较长的序列，建议在内部连接FRET对。

下图是武汉天德生物多肽荧光修饰案例之一：

多肽荧光修饰

FRET对

荧光共振能量转移（FRET）是一种描述两种荧光团之间能量转移的机制。由于FRET效率部分取决于供体分子和受体分子之间的距离，因此该技术通常用于研究酶的效率、蛋白质与蛋白质之间相互作用或其他分子动力学（图1）。

用于亚细胞成像的荧光多肽

利用共焦显微镜或荧光显微镜进行体外成像仍然是研究细胞内各种生物学过程和相互作用的最高效、最有效的方法之一。当与成像技术结合时，荧光标记多肽可用于识别特定靶标。

For example, cell penetrating peptides (CPPs) modified with FITC and photoactivatable probes have been used to track binding patterns and dynamic behavior over time. Unlike proteins, these peptides localize to specific targets on actin and are less prone to protein aggregation, making them ideal for in vitro tracking (Pan 2014). Similarly, FITC-labeled CPPs have been used to image intracellular compartments of cells with low risk of cytotoxicity (Kirkham 2015).

用于血管造影的荧光多肽

用于血管造影的体内成像仍然是荧光标记多肽的主要应用领域。该技术使用造影剂对血管内部进行成像，使医生能够为个人选择最合适的医疗策略或治疗方法。

For example, high resolution near-infrared fluorophores have been used as fibrin imaging-agents for deep vein thrombosis (DVT). Using Cy7 labeled fibrin-targeting peptides, CT scanning and confocal microscopy were used to differentiate between acute and subacute murine DVT (Hara 2012). Similar techniques have also been used for detecting apoptosis as a symptom for glaucoma. Fluorophore-labeled peptides that are activated by caspases can be imaged in vivo to track glaucoma progression (Qiu 2014).

用于筛选蛋白水解肽的FRET

蛋白水解酶在传染性疾病中发挥重要作用，使其成为开发新疗法的研究目标。为了识别这些酶靶向的肽序列，通常使用肽文库。潜在的蛋白酶靶序列与FRET对结合在一起，这样当酶裂解靶肽时，可以检测到荧光信号。

In these studies, a donor molecule, such as Abz (Marcondes 2015) or Lucifer Yellow (Rossé 2000) is covalently attached to the C-terminus, while acceptor molecules (ex. Dabsyl, DNP) are coupled to the N-terminus. This strategy is commonly employed with peptides synthesized using a solid-phase approach, and peptides are left conjugated to the bead. If the peptides are not targeted by protease activity, both the donor and acceptor will be present, and the bead will appear non-fluorescent. If the peptides are cleaved, the peptide will no longer be quenched by FRET and the beads will fluoresce. Since protease studies are most effective when FRET is combined with shorter peptides, micro-scale peptide libraries are ideal choices for these experiments.