Fluorescein isothiocyanate (FITC) is a widely used due to its high quantum efficiency and stability when conjugated. FITC exhibits a yellow-orange hue, characterized by its absorption peak at 495 nm. Upon excitation, it emits a yellow-green fluorescence, with its emission peak centered at 525 nm. The conjugation of FITC to antibodies represents an invaluable technique, offering the capacity to permanently tag biomolecules with a distinctive detectable property. This feature serves as a powerful tool for the visualization and tracking of the conjugate's interactions with other biomolecules. FITC's application extends to the identification of surface molecules, facilitating investigations through fluorescence microscopy or flow cytometry. Presented below is a detailed procedure outlining the coupling of antibody molecules with fluorescein derivatives.
FITC, derived from the nature fluorescein molecule, is characterized by the introduction of an isothiocyanate reactive group (−N=C=S), which replaces a hydrogen atom located on the lower ring of its molecular structure. The synthesis of FITC primarily involves the modification of the lower ring, specifically at the 5-carbon positions. This unique isothiocyanate functionality within FITC enables it to interact with primary amine groups found in proteins or antibodies, rendering it a pivotal tool for biomolecular labeling purposes.
Fig. 1 The molecular architecture of FITC.
Fig. 2 The pipeline of labeling antibody with FITC.
PDCD1 HIV1 gp120 CD33 IAV HA PRNP TNF MS4A1 CD274 APP EGFR CD40 ERBB3 TFPI ERBB2 SARS-CoV S PCSK9 MET BSG HAVCR2 IL6 CD3 LAG3 ROR1 TNFRSF4 IL33 CD19 CD38 JAG1 IL17A IFNG CD47 HCV E2 Protein IL1A / IL1B HBsAg IGF1R CD4 AXL ANGPT2 IL23A NGF MUC1 TNFRSF18 TNFRSF9 MSTN IL1B HRSV F Protein KDR SORT1 C5 VEGF CXCR4 TNFRSF17 Tau TFRC CTLA4 IL13 IAV H5N1 HA TSLP MSLN SOST
Fig. 3 Flow cytometry-based analysis of CD19 B-lymphocytic antigen in human donor lymphocytes.1
Two categories of FITC-labeled antibodies were employed in the labeling process of human leukemic cells: FITC-conjugated mouse antibodies targeting Human CD19 protein (referred to as anti-CD19-FITC*) and FITC-conjugated mouse antibodies against Human IgG1 protein (designated as anti-IgG1-FITC*).
Fig. 4 Microscopic visualization of FITC-conjugated anti-SAG1 IgY antibody-stained T. gondii tachyzoites.2
This investigation marks a pioneering achievement in the development of a FITC-conjugated anti-SAG1 IgY antibody, as substantiated by ELISA, fluorescence spectrometry, and cell culture fluorescence imaging. The FITC-labeled IgY antibodies specific to T. gondii tachyzoites, thus obtained, hold promise for future deployment as diagnostic instruments for detecting T. gondii infections across diverse sample types.
Creative Biolabs offers antibody labeling services using FITC. For additional details, feel free to contact us without hesitation.
