Chemigenetic Calcium Indicators Brightness Photostability
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Solution Overview
Problem
Existing genetically encoded fluorescent calcium indicators (GECIs) have limitations in brightness, photostability, and spectral range, which hinder their effectiveness in imaging neuronal activity.
Innovation Solution
Development of chemigenetic calcium indicators that combine a calcium-binding protein domain with a ligand binding protein domain, such as HaloTag, and a dye-ligand conjugate, enhancing brightness, photostability, and spectral range.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Illumination intensity
If genetically encoded fluorescent calcium indicators (GECIs) are used, then calcium imaging capability is provided, but brightness and photostability are limited
Solution Approach 1:
The patent combines a calcium-binding protein domain (genetic component) with a small-molecule fluorescent dye (synthetic component) to create a hybrid chemigenetic calcium indicator. This composite structure integrates the calcium selectivity and biological compatibility of GECIs with the superior brightness and photostability of synthetic dyes, directly resolving the contradiction between imaging capability and optical performance.
2Adaptability or versatility
If fluorescent protein domains are used in GECIs, then calcium-binding functionality is achieved, but spectral range is limited
Solution Approach 1:
The patent divides the calcium indicator into two functional segments: a calcium-binding protein domain that maintains calcium selectivity and a small-molecule fluorescent dye that provides enhanced spectral properties. This segmentation allows each component to optimize its specialized function while working together as an integrated system.
Solution Approach 2:
The patent changes the optical parameters of the indicator by replacing the fluorescent protein chromophore with a small-molecule fluorescent dye, thereby expanding the spectral range while preserving the calcium-binding functionality through the protein domain.
3Reliability
If small molecule synthetic fluorophores are used, then brightness and photostability are improved, but calcium-selective binding is lost
Solution Approach 1:
The patent merges the calcium-binding capability of protein domains with the superior optical properties of small-molecule fluorescent dyes through a covalent linkage, creating a unified chemigenetic indicator that exhibits both calcium selectivity and enhanced photostability simultaneously.
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 chemigenetic calcium indicators provide improved brightness, photostability, and spectral range compared to traditional fluorescent proteins, enabling more effective imaging of neuronal activity.
Implementation Method 1
determining changes in fluorescence, the chemigenetic calcium indicator comprising a ligand binding protein domain having a calcium-binding protein domain and a dye-ligand conjugate attached thereto
Data Source
AI summary
A chemigenetic calcium indicator and a method of measuring calcium are provided. The chemigenetic calcium indicator includes a calcium-binding protein domain attached to a ligand binding protein domain. The method of measuring calcium includes administering a chemigenetic calcium indicator to a subject and determining changes in fluorescence, the chemigenetic calcium indicator including a ligand binding protein domain having a calcium-binding protein domain and a dye-ligand conjugate attached thereto.