Fluorescence Labeling of Peptides

As a specialized provider of biochemical services, BOC Sciences offers service for fluorescent labeling of peptides. We provide a wide range of fluorescent dyes for peptide labeling, such as Fluorescein isothiocyanate, Carboxyfluorescein, Carboxytetramethylrhodamine, AMC, ATTO, Alexa Fluor, Bodipy, Cy3, Cy5...

Peptide Fluorescent Labeling

Fluorescent labeling technology refers to the use of a fluorescent substance covalently bound or physically adsorbed to a group of molecules to be studied, using its fluorescent properties to provide information about the object under study. Researchers can use fluorescently labeled peptides to detect the activity of target proteins and apply the high-throughput activity screening methods they have developed to drug screening and drug development for disease therapeutic target proteins.

Fig.1 Fluorophores and strategies for fluorescent peptide labeling. (Mendive‐Tapia et al., 2021)

Fluorescent Labeling of Peptides Services

BOC Sciences' services are designed to bind fluorescent dyes to peptide molecules to generate fluorescently labeled peptide compounds.

FITC Modified Peptides

Fluorescein isothiocyanate (FITC) is highly reactive, and it is generally easier to be introduced into a solid phase synthesis than other fluoresceins, and the reaction does not require the addition of activation reagents. BOC Sciences synthesizes FITC-modified peptides in two general forms.

Mode 1: FITC is inserted at the end of the entire peptide chain and a molecule of Acp (6-aminohexanoic acid) is inserted before the FITC. During the reaction, the access of Acp provides a straight chain space of six carbons, which greatly reduces the spatial site resistance of the reaction, improves the reaction efficiency, and reduces the difficulty of the reaction.

Mode 2: FITC is inserted into a certain Lys side chain in the whole peptide, and the Lys side chain is a four-carbon straight-chain alkyl group with -NH₂ at the end, which directly plays a role in lowering the spatial site resistance.

AMC Modified Peptides

7-Amino-4-methylcoumarin (AMC) is a widely used fluorescent labeling reagent that modifies peptide molecules from the C-terminus in the following steps.

1. AMC reacts with the first amino acid at the C-terminus of the peptide chain.
2. Solid-phase synthesis of the entire peptide chain (starting from the second amino acid) and retaining the side-chain protecting groups and the last amino protecting group of the entire peptide chain.
3. Liquid phase condensation of AA-AMC with the fully protected peptide chain.
4. Excision of the protecting group completes the modification of the peptide chain.

In addition to the above fluorescent substances, we can also provide 5-FAM, Rhodamine B, 5-TAMRA, Cy 3, Cy 5, and other substances for peptide labeling.

Peptide Fluorescent Labeling Test

• Gel Electrophoresis Analysis: By comparing a labeled peptide with an unlabeled peptide, the difference in migration between the two on the gel can be observed, and the efficiency of the labeling can be calculated.
• High-Performance Liquid Chromatography (HPLC) Analysis: By detecting the absorption or emission signals of fluorescent dyes, the amount of labeled peptide can be quantitatively analyzed and the efficiency of labeling can be evaluated.
• Functionality Testing: Based on the functional properties of the peptides, corresponding functional tests can be designed to assess whether the activity and function of the labeled peptide are affected by the labeling.

Frequently Asked Questions (FAQ)

How does fluorescent labeling of peptides help in protein-protein interaction studies?

Fluorescently labeled peptides allow real-time monitoring of protein-protein interactions by tracking the fluorescence signal. This method provides insights into the kinetics and dynamics of these interactions, aiding in high-throughput screening and structural studies.

Can I use multiple fluorescent labels on a single peptide?

Yes, multiple fluorescent labels can be attached to a single peptide, allowing for multiplexed detection and analysis. This approach can help track several molecular processes simultaneously, increasing the depth of your experimental data.

What are the key differences between using FITC and AMC for peptide labeling?

FITC provides brighter fluorescence and is ideal for studies that require high sensitivity, whereas AMC is useful for assays involving peptide cleavage or enzyme activity because it emits fluorescence upon cleavage. The choice depends on the specific experimental needs and sensitivity requirements.

How do I determine the optimal concentration of fluorescent dye for my peptide labeling?

The optimal concentration of fluorescent dye depends on factors like the peptide's size, the type of dye, and the specific application. We recommend conducting preliminary trials to determine the optimal labeling efficiency and fluorescence intensity for your experiment.