Spectrofluorometer

A spectrofluorometer is an analytical instrument that measures fluorescence, allowing researchers to detect and quantify fluorescent molecules in biological and chemical samples. It uses a monochromator or selected light source to excite fluorophores at specific wavelengths, then separates and records the longer-wavelength emission produced as excited molecules return to a lower-energy state. In immunology and infection research, spectrofluorometers support assays involving fluorescently labeled antibodies, antigen-antibody interactions, enzyme activity, and pathogen-associated molecules. By converting fluorescence intensity into quantitative measurements, they help characterize immune responses, evaluate diagnostic assays, and monitor molecular changes during infection.

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Research

JoVE EoE - Bacterial Growth and Techniques

Quantifying Bacterial Infection-Induced Amyloids Using Thioflavin T Fluorescence

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2025

Source: Balczon, R., et al. Methods for Detecting Cytotoxic Amyloids Following Infection of Pulmonary Endothelial Cells by Pseudomonas aeruginosa. J. Vis. Exp. (2018)This video demonstrates the protocol to quantify amyloids in culture supernatants from pulmonary endothelial cells infected with virulent bacteria, using thioflavin T, a fluorescent dye that binds to amyloid beta-sheet structures. An increase in fluorescence compared to baseline indicates the presence of amyloids in the culture...

Studying the Permeation of FITC and FITC-Ferritin through an In Vitro Blood-Brain Barrier Model

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2025

This video demonstrates a method to study permeation through an in vitro blood-brain barrier (BBB) model using astrocytes and endothelial cells on a membrane insert, with free FITC and FITC-loaded ferritin. FITC-ferritin bound to receptors on endothelial cells and crossed the membrane barrier to the lower chamber while free FITC showed restricted permeation and remained in the upper chamber.

Fluorescence Leakage Assay to Study Cell-Penetrating Peptide Interaction with Membrane

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2025

This video discusses the fluorescence leakage assay to study the interactions of cell-penetrating peptides with cell membranes using cell membrane mimics — large unilamellar vesicles, or LUVs, encapsulated with a fluorescent dye and a quencher.

Fluorescence Anisotropy-Based Detection of Protein-Protein Interactions

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2025

In this video, we describe the fluorescence anisotropy technique to study the interactions between the fluorophore-tagged Shwachman-diamond syndrome (SBDS) protein and the elongation factor-like 1 GTPase (EFL1). On incubating SBDS proteins with gradually increasing concentrations of EFL1, a steady increase in anisotropy is observed, indicating a successful interaction between the two proteins.

In Vitro Permeation of FITC-loaded Ferritins Across a Rat Blood-brain Barrier: a Model to Study the Delivery of Nanoformulated Molecules

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Cited by 15 •

2016

A method to establish an in vitro model of blood-brain barrier based on a co-culture of rat brain microvascular endothelial cells and astrocytes is described and validated. This system proved to be a valid tool to study the effect of nanoformulation on the trans-barrier permeation of fluorescent molecules.

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