Synthesis, Cellular Delivery and <em>In vivo</em> Application of Dendrimer-based pH Sensors

Lorenzo Albertazzi; Barbara Storti; Marco Brondi; Sebastian Sulis Sato; Gian Michele Ratto; Giovanni Signore; Fabio Beltram

doi:10.3791/50545

Synthesis, Cellular Delivery and In vivo Application of Dendrimer-based pH Sensors

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Chemistry

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JoVE Journal Chemistry

Synthesis, Cellular Delivery and In vivo Application of Dendrimer-based pH Sensors

Synthesis, Cellular Delivery and In vivo Application of Dendrimer-based pH Sensors

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16:19 min

September 10, 2013

DOI: 10.3791/50545-v

Lorenzo Albertazzi¹, Barbara Storti², Marco Brondi², Sebastian Sulis Sato², Gian Michele Ratto³, Giovanni Signore⁴, Fabio Beltram²

¹Institute for Complex Molecular Systems & Laboratory of Macromolecular and Organic Chemistry,Eindhoven University of Technology & NEST, Scuola Normale Superiore and Istituto Nanoscienze-CNR, ²NEST, Scuola Normale Superiore and Istituto Nanoscienze-CNR & IIT@NEST,Center for Nanotechnology Innovation, ³NEST, Scuola Normale Superiore and Istituto Nanoscienze-CNR, ⁴IIT@NEST,Center for Nanotechnology Innovation

Overview

This article presents a methodology for synthesizing dendrimer-based fluorescent sensors aimed at measuring pH levels in living cells and in vivo. The use of a dendritic scaffold enhances the properties of conjugated fluorescent dyes, resulting in improved sensing capabilities.

Key Study Components

Area of Science

Fluorescence imaging
Cellular biology
Neuroscience

Background

Fluorescence sensors are essential tools in life sciences.
Dendrimers can improve the performance of fluorescent dyes.
pH measurement is crucial for understanding cellular environments.
Existing methods may lack sensitivity or specificity.

Purpose of Study

To develop new fluorescent sensors for pH imaging.
To enhance the properties of pH-sensitive dyes using dendritic scaffolds.
To enable in vitro and in vivo pH measurements in living cells.

Methods Used

Conjugation of pH-sensitive dyes to dendritic scaffolds.
In vitro measurements to create a pH calibration curve.
Electroporation for sensor delivery into cells.
Confocal microscopy for imaging pH in the extracellular space.

Main Results

Successful synthesis of dendrimer-based fluorescent sensors.
Improved sensitivity and specificity in pH measurements.
Effective imaging of pH in living cells and brain tissue.
Demonstrated utility during physiological and pathological conditions.

Conclusions

Dendrimer-based sensors represent a significant advancement in pH imaging.
These sensors can be applied in various biological contexts.
Future studies may explore additional applications in neuroscience.

Frequently Asked Questions

What are dendrimer-based fluorescent sensors?

Dendrimer-based fluorescent sensors are advanced tools that utilize a dendritic scaffold to enhance the properties of fluorescent dyes for improved sensing capabilities.

How are these sensors delivered into cells?

The sensors are delivered using electroporation, a technique that facilitates the introduction of molecules into cells through electrical pulses.

What imaging techniques are used in this study?

Confocal microscopy and two-photon microscopy are employed to visualize pH levels in living cells and brain tissue.

What is the significance of measuring pH in living cells?

Measuring pH in living cells is crucial for understanding cellular processes and can provide insights into physiological and pathological conditions.

Can these sensors be used in vivo?

Yes, the study demonstrates the application of these sensors for in vivo pH imaging in the brain of anesthetized mice.

Fluorescence sensors are powerful tools in life science. Here we describe a methodology to synthesize and use dendrimer-based fluorescent sensors to measure pH in living cells and in vivo. The dendritic scaffold enhances the properties of conjugated fluorescent dyes leading to improved sensing properties.

The overall goal of the following experiment is to develop new fluorescent sensors based on Dun drummers with improved properties for pH imaging in vitro in living cells and in vivo. This is achieved by first conjugating pH sensitive dyes to a dendritic scaffold together with other moieties, such as targeting groups or pH in sensitive dyes. And in vitro measurements are taken to generate a pH calibration curve next to image pH in living cells.

The sensor is delivered by electroporation and pH maps are generated using confocal microscopy to image pH in the extracellular space of the brain. During physiological and pathological neural activity, the sensor is micro injected into the brain of anesthetized mice. Fluorescence data is then acquired using two photon microscopy.

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