Synthesis and Identification of Biologically Active Mono-Labelled FITC-Insulin Conjugate.

Area of Science:

• Biochemistry
• Protein Chemistry
• Pharmaceutical Research

Background:

• Fluorescently labelling proteins like insulin is crucial for pharmaceutical research and drug delivery.
• Understanding protein conjugation is key to developing new therapeutic and diagnostic tools.

Purpose of the Study:

• To synthesize and characterize fluorescein isothiocyanate (FITC)-labelled human insulin.
• To evaluate the biological activity of FITC-insulin conjugates compared to native insulin.
• To explore the potential of FITC-insulin in biomedical applications.

Main Methods:

• Human insulin (Actrapid®) was labelled with FITC using varying reaction times and pH.
• Conjugates were identified using reverse-phase high-performance liquid chromatography (RP-HPLC) and electrospray mass spectroscopy.
• Biological activity was assessed by examining AKT phosphorylation in HUVEC cells and GLUT4 expression in C2C12 cells.

Main Results:

• Mono-labelled FITC-insulin at the B1 position was successfully synthesized with high yield.
• FITC-insulin conjugates retained significant biological activity, with monoB1 showing 99.5% activity.
• MonoB1 FITC-insulin demonstrated comparable cellular response to native insulin in C2C12 skeletal muscle cells.

Conclusions:

• Mono-labelled FITC-insulin at the B1 position exhibits similar biological activity to native insulin.
• This conjugate holds promise for future biomedical applications, including drug delivery and research.
• The study validates FITC-insulin as a viable tool for investigating insulin's biological functions.