Source 1primary paper2025MED
Toolkit/CheRiff + jRCaMP1b + RH237 cardiac all-optical electrophysiology platform
CheRiff + jRCaMP1b + RH237 cardiac all-optical electrophysiology platform
Also known as: experimental platform, optogenetic stimulation with simultaneous Vm and CaT imaging
Taxonomy: Mechanism Branch / Architecture. Workflows sit above the mechanism and technique branches rather than replacing them.
Summary
We developed a novel experimental platform in which tissue is stimulated optogenetically while Vm and CaT are imaged simultaneously.
Usefulness & Problems
Why this is useful
This platform optogenetically stimulates engineered cardiac spheroids while simultaneously imaging membrane potential and calcium transients. It uses CheRiff for actuation, jRCaMP1b for calcium reporting, and RH237 for voltage reporting.; simultaneous optogenetic stimulation and dual optical readout of membrane potential and calcium transients; studying coupling between engineered tissue grafts and host tissue
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This platform optogenetically stimulates engineered cardiac spheroids while simultaneously imaging membrane potential and calcium transients. It uses CheRiff for actuation, jRCaMP1b for calcium reporting, and RH237 for voltage reporting.
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simultaneous optogenetic stimulation and dual optical readout of membrane potential and calcium transients
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studying coupling between engineered tissue grafts and host tissue
Problem solved
It provides simultaneous stimulation and dual functional optical mapping in engineered cardiac tissue. The abstract also states it is well-suited for distinguishing graft versus host activation in coupling studies.; combines stimulation with simultaneous Vm and CaT imaging in engineered cardiac spheroids; enables independent stimulation and attribution of activation to graft or host tissue
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It provides simultaneous stimulation and dual functional optical mapping in engineered cardiac tissue. The abstract also states it is well-suited for distinguishing graft versus host activation in coupling studies.
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combines stimulation with simultaneous Vm and CaT imaging in engineered cardiac spheroids
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enables independent stimulation and attribution of activation to graft or host tissue
Problem links
combines stimulation with simultaneous Vm and CaT imaging in engineered cardiac spheroids
LiteratureIt provides simultaneous stimulation and dual functional optical mapping in engineered cardiac tissue. The abstract also states it is well-suited for distinguishing graft versus host activation in coupling studies.
Source:
It provides simultaneous stimulation and dual functional optical mapping in engineered cardiac tissue. The abstract also states it is well-suited for distinguishing graft versus host activation in coupling studies.
enables independent stimulation and attribution of activation to graft or host tissue
LiteratureIt provides simultaneous stimulation and dual functional optical mapping in engineered cardiac tissue. The abstract also states it is well-suited for distinguishing graft versus host activation in coupling studies.
Source:
It provides simultaneous stimulation and dual functional optical mapping in engineered cardiac tissue. The abstract also states it is well-suited for distinguishing graft versus host activation in coupling studies.
Published Workflows
Objective: Develop an all-optical cardiac electrophysiology platform that enables optogenetic stimulation together with simultaneous membrane-potential and calcium-transient imaging in human engineered cardiac spheroids.
Why it works: The platform uses blue light to excite the optogenetic actuator and a single green band to excite both optical reporters, then separates emitted fluorescence into calcium and voltage channels on two cameras to avoid cross-talk.
optogenetic actuation with CheRiffgenetically encoded calcium reporting with jRCaMP1bvoltage reporting with RH237spectral separation of excitation and emission channelsoptogenetic stimulationsimultaneous dual-channel optical mappingsplit-emission two-camera imagingcomparison of alternative green excitation bands
Stages
- 1.Engineer and prepare cardiac spheroids with actuator and reporters(library_build)
This stage establishes the engineered tissue and optical components required for simultaneous stimulation and dual readout.
Selection: Prepare model tissue containing human iPSC-derived cardiomyocytes and fibroblasts with genetically encoded CheRiff and jRCaMP1b plus RH237 voltage dye.
- 2.Configure excitation and split-emission imaging(functional_characterization)
This stage implements the optical design needed to collect simultaneous calcium and voltage signals while minimizing channel interference.
Selection: Use blue light for CheRiff excitation and one of two green bands for RH237 and jRCaMP1b excitation, with fluorescence split to two cameras.
- 3.Test stimulation and compare excitation bands(confirmatory_validation)
This stage confirms that the platform works in the target tissue and identifies the trade-off between signal quality and perturbation from the excitation band choice.
Selection: Assess successful optogenetic stimulation, simultaneous Vm and CaT recording without cross-talk, and compare signal-to-noise and excitability effects between the two green excitation bands.
Steps
- 1.Generate human engineered cardiac spheroids containing iPSC-derived cardiomyocytes and fibroblasts
Establish the model tissue used for the all-optical platform.
The tissue model must exist before genetic encoding, dye staining, stimulation, and imaging can be performed.
- 2.Genetically encode spheroids with CheRiff and jRCaMP1bactuator and calcium reporter
Install optogenetic actuation and genetically encoded calcium sensing in the tissue.
Genetic encoding is required before the optical stimulation and calcium-imaging functions can be tested.
- 3.Stain spheroids with RH237 voltage dyevoltage reporter
Enable membrane-potential imaging alongside calcium imaging.
Voltage dye staining is needed before simultaneous Vm and CaT optical mapping can be performed.
- 4.Excite CheRiff with blue light and excite RH237 and jRCaMP1b with a single green bandactuator and reporters under optical excitation
Drive optogenetic stimulation while collecting both reporter signals under a compatible excitation scheme.
The excitation configuration must be set before simultaneous imaging can be evaluated.
- 5.Split fluorescence emission and image calcium and voltage channels on two cameras
Separate CaT and Vm signals for simultaneous acquisition with minimal cross-talk.
Emission splitting follows excitation and is the core measurement step that enables independent readout of the two signals.
- 6.Compare the two green excitation bands for cross-talk, signal-to-noise, and excitability effectsplatform under comparative evaluation
Determine the trade-off between signal quality and unintended CheRiff activation across excitation-band choices.
Band comparison is meaningful only after the full stimulation and imaging setup is operating.
Taxonomy & Function
Primary hierarchy
Mechanism Branch
Architecture: A reusable architecture pattern for arranging parts into an engineered system.
Mechanisms
fluorescence-based calcium reportingoptogenetic depolarizationsimultaneous dual-parameter optical imagingvoltage-sensitive dye reportingTechniques
No technique tags yet.
Target processes
recombinationInput: Light
Implementation Constraints
cofactor dependency: cofactor requirement unknownencoding mode: genetically encodedimplementation constraint: context specific validationimplementation constraint: spectral hardware requirementoperating role: sensorswitch architecture: split
The source describes human iPSC-derived cardiac spheroids genetically encoded with CheRiff and jRCaMP1b, stained with RH237, excited with blue and green light, and imaged with split emission on two cameras.; requires genetic encoding of CheRiff and jRCaMP1b in the tissue construct; requires RH237 staining; requires blue-light excitation for CheRiff and green-light excitation with split-emission imaging on two cameras
The abstract does not show that the platform avoids all perturbation, because one excitation band slightly increased tissue excitability through CheRiff activation.; the 525-575 nm excitation band caused a slight increase in tissue excitability because of CheRiff activation
Validation
Cell-freeBacteriaMammalianMouseHumanTherapeuticIndep. Replication
Observations
successMammalian Cell Lineapplication demohuman
simultaneous Vm and CaT optical mapping
Inferred from claim claim3 during normalization. The platform enabled successful optogenetic stimulation and simultaneous recording of membrane potential and calcium transients without cross-talk using either 525-575 nm or 558-575 nm green excitation. Derived from claim claim3.
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Supporting Sources
Ranked Claims
The 525-575 nm excitation band produced higher signal-to-noise ratios than the 558-575 nm band but slightly increased tissue excitability because of CheRiff activation.
Human engineered cardiac spheroids were genetically encoded with CheRiff and jRCaMP1b and stained with RH237 for simultaneous all-optical electrophysiology.
The platform enabled successful optogenetic stimulation and simultaneous recording of membrane potential and calcium transients without cross-talk using either 525-575 nm or 558-575 nm green excitation.
The authors developed an experimental platform that combines optogenetic stimulation with simultaneous optical imaging of membrane potential and calcium transients.
The system is well-suited for studying coupling between engineered tissue grafts and host tissue because the two tissue types can be stimulated independently and activation can be attributed to graft or host.
Approval Evidence
1 source5 linked approval claimsfirst-pass slug cheriff-jrcamp1b-rh237-cardiac-all-optical-electrophysiology-platform
We developed a novel experimental platform in which tissue is stimulated optogenetically while Vm and CaT are imaged simultaneously.
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comparative performancesupports
The 525-575 nm excitation band produced higher signal-to-noise ratios than the 558-575 nm band but slightly increased tissue excitability because of CheRiff activation.
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component usagesupports
Human engineered cardiac spheroids were genetically encoded with CheRiff and jRCaMP1b and stained with RH237 for simultaneous all-optical electrophysiology.
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performancesupports
The platform enabled successful optogenetic stimulation and simultaneous recording of membrane potential and calcium transients without cross-talk using either 525-575 nm or 558-575 nm green excitation.
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platform capabilitysupports
The authors developed an experimental platform that combines optogenetic stimulation with simultaneous optical imaging of membrane potential and calcium transients.
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use casesupports
The system is well-suited for studying coupling between engineered tissue grafts and host tissue because the two tissue types can be stimulated independently and activation can be attributed to graft or host.
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Comparisons
Source-stated alternatives
Within this paper, the main alternative configuration is using the 558-575 nm green excitation band instead of 525-575 nm, trading lower signal-to-noise for less CheRiff-related excitability increase.
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Within this paper, the main alternative configuration is using the 558-575 nm green excitation band instead of 525-575 nm, trading lower signal-to-noise for less CheRiff-related excitability increase.
Source-backed strengths
simultaneous Vm and CaT recording without cross-talk using both tested excitation bands; combines genetically encoded actuator and calcium indicator with a voltage dye that can stain any tissue
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simultaneous Vm and CaT recording without cross-talk using both tested excitation bands
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combines genetically encoded actuator and calcium indicator with a voltage dye that can stain any tissue
Compared with CheRiff
Within this paper, the main alternative configuration is using the 558-575 nm green excitation band instead of 525-575 nm, trading lower signal-to-noise for less CheRiff-related excitability increase.
Shared frame: source-stated alternative in extracted literature
Strengths here: simultaneous Vm and CaT recording without cross-talk using both tested excitation bands; combines genetically encoded actuator and calcium indicator with a voltage dye that can stain any tissue.
Relative tradeoffs: the 525-575 nm excitation band caused a slight increase in tissue excitability because of CheRiff activation.
Source:
Within this paper, the main alternative configuration is using the 558-575 nm green excitation band instead of 525-575 nm, trading lower signal-to-noise for less CheRiff-related excitability increase.
Ranked Citations
- 1.