Olfactory CNG Channel Cell Assay Automation
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Solution Overview
Problem
Current methods for high-throughput screening of olfactory CNG channel modulators are limited by the need for electrophysiological assays, which are not amenable to automation, and there is a lack of identified human olfactory CNG channel subunits for targeted modulation of smell perception.
Innovation Solution
The development of isolated nucleic acid sequences encoding human OCNC1, OCNC2, and β1b subunits of the olfactory CNG channel, along with methods for expressing these in host cells, such as HEK-293 cells, to create cell-based assays using fluorescent calcium or membrane potential dyes for high-throughput screening of compounds that modulate olfactory CNG channel activity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If electrophysiological assays are used to screen olfactory CNG channel modulators, then measurement precision is improved, but ease of operation and extent of automation deteriorate
Solution Approach 1:
The patent replaces electrophysiological measurement methods with optical detection methods. Specifically, it uses fluorescent dyes (such as fluorescein diacetate) that change fluorescence properties in response to ion flux through CNG channels, substituting electrical measurement systems with optical detection systems that are more amenable to high-throughput automation while maintaining detection sensitivity
2Productivity
If human olfactory CNG channel subunits are identified and expressed in host cells, then productivity is improved, but device complexity increases
Solution Approach 1:
The patent creates a universal heterologous expression system where human olfactory CNG channel subunits (OCNC1, OCNC2, and beta1b) are expressed in common host cells (such as HEK-293 or CHO cells). This universal system can screen multiple compounds and channel variants using the same basic platform, reducing overall complexity despite the added capability of expressing specific human subunits
Solution Approach 2:
The patent introduces cDNA encoding human CNG channel subunits as intermediaries between the host cell's endogenous machinery and the desired channel function. These cDNAs serve as mediators that allow the host cell's transcription-translation apparatus to produce functional human CNG channels, enabling high-throughput screening without requiring complex direct manipulation of channel proteins
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
Enables the identification of compounds that enhance or inhibit smell by modulating olfactory CNG channel activity, facilitating the development of smell-enhancing or smell-blocking agents and providing a platform for screening modulators of G-protein coupled receptors and other proteins that regulate cyclic nucleotide levels.
Implementation Method 1
using fluorescent calcium or membrane potential dyes for high-throughput screening
Implementation Method 2
using fluorescent calcium or membrane potential dyes for high-throughput screening
Data Source
AI summary
The present invention relates to isolated nucleic acid sequences that encode human olfactory cyclic nucleotide gated (CNG) channel subunits, and the corresponding polypeptides. The invention further relates to the use of human CNG channels to profile, screen for, and identify compounds that modulate the human olfactory CNG channel. More specifically, the invention relates to the expression of the human olfactory CNG channel in cells, preferably mammalian cells, and the use of these cells in high throughput cell-based assays to identify compounds that enhance or block human olfactory CNG function. Compounds that activate the olfactory CNG channel will enhance smell and can be used to make foods more palatable for individuals with attenuated olfactory function. Conversely, compounds that inhibit the olfactory CNG channel will inhibit smell and can be use to block malodors. Additionally, the invention relates to the use of cell-based olfactory CNG channel assays to identify modulates of G-protein coupled receptor (GPCRs) and other proteins that regulate cyclic nucleotide levels.