Cyanine Dye Composition for Multichannel Live-Cell Staining
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Solution Overview
Problem
Existing methods for in vivo staining of microorganisms and living cells using GFP are hindered by antibiotic resistance, and conventional cyanine dyes interfere with cell viability, limiting multichannel fluorescence observation.
Innovation Solution
Development of cyanine dyes that bind to nucleic acid and exhibit different fluorescence colors without significantly reducing cell viability, allowing multichannel observation through simultaneous staining of cells with multiple dyes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional cyanine dyes are used for in vivo staining, then cells can be stained with fluorescent dyes, but cell viability is significantly reduced
Solution Approach 1:
The patent modifies the chemical structure of cyanine dyes by changing parameters such as the sulfur atom substitution (replacing with oxygen or nitrogen), modifications to the heterocyclic rings, and adjustments to substituent groups. These parameter changes result in dyes with reduced toxicity while maintaining fluorescence properties, thereby resolving the contradiction between staining capability and cell viability.
Solution Approach 2:
The invention creates composite dye molecules combining modified cyanine structures with specific substituents (such as triphenylphosphine, carboxylic acid groups, or hydroxyl groups). These composite structures provide both the necessary fluorescence for staining and reduced cytotoxicity, simultaneously achieving measurement precision and reliability.
2Measurement precision
If GFP expression is used to investigate microorganisms, then fluorescent labeling is achieved, but antibiotic resistance prevents transformation selection
Solution Approach 1:
The patent extracts the fluorescent labeling function from the GFP protein system and implements it through small molecule cyanine dyes that can permeate cell membranes and bind to nucleic acids. This extraction allows fluorescent labeling without requiring genetic transformation, thereby overcoming the limitation of antibiotic resistance in drug-resistant bacteria.
Solution Approach 2:
The cyanine dyes act as intermediaries that provide fluorescent labeling capability without requiring direct genetic manipulation. The dyes penetrate cells and bind to nucleic acids, serving as a mediator between the observer and the microorganism, enabling investigation of drug-resistant strains that cannot be genetically transformed.
3Adaptability or versatility
If multiple cyanine dyes are used for multichannel staining, then different cell populations can be observed, but dye toxicity increases
Solution Approach 1:
The patent develops a series of cyanine dyes with different fluorescence emission wavelengths by modifying chemical parameters such as the number of methine groups, heterocyclic ring substitutions, and auxiliary groups. These parameter variations enable multichannel staining while maintaining low toxicity across all dye variants, allowing simultaneous observation of multiple cell populations.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The new cyanine dyes maintain cell viability, enabling effective multichannel fluorescence microscopy for observing interactions among different cell populations.
Implementation Method 1
cyanine dyes which are not reducing the viability of the stained cells substantially, in particular cyanine dyes binding to nucleic acid
Implementation Method 2
exerting red, green and yellow fluorescence upon excitation with light of suitable wavelength
Data Source
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AI summary
The invention provides new cyanine dyes for staining living cells, providing fluorescence emission in red, green and yellow, thus allowing "multichannel" staining. The dyes are binding to nucleic acids and allow the observation of the stained cells, for the staining does not negatively interfere with the viability of the stained cells. The inventive dyes thus can advantageously be used for pathogen-host investigations or any other type of investigation of cell-cell-interactions, for the natural behavior of the stained cells (microorganisms, pathogens etc.) can easily be observed.