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Related Experiment Video

Updated: Apr 20, 2026

Direct Imaging of ER Calcium with Targeted-Esterase Induced Dye Loading TED
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Improving AM ester calcium dye loading efficiency.

Mohammad I K Hamad1, Martin Krause2, Petra Wahle1

  • 1AG Entwicklungsneurobiologie, Fakultät für Biologie, Ruhr Universität Bochum, D-44780 Bochum, Germany.

Journal of Neuroscience Methods
|December 3, 2014
PubMed
Summary
This summary is machine-generated.

Optimizing acetoxymethyl ester (AM) dye loading for calcium imaging in slice cultures requires lower concentrations of Pluronic F127 (PF127) and dimethyl sulfoxide (DMSO). This improved method enhances staining efficiency and preserves neuronal function for long-term imaging.

Keywords:
AM ester dyeCalcium imagingDMSOOrganotypic culturesOsmolalityPluronic PF127

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Area of Science:

  • Neuroscience
  • Cellular Biology
  • Biochemistry

Background:

  • Calcium imaging is crucial for understanding neuronal function.
  • Acetoxymethyl ester (AM) dyes are commonly used but their loading efficiency is affected by Pluronic F127 (PF127) and dimethyl sulfoxide (DMSO) concentrations.
  • Optimal concentrations of PF127 and DMSO for AM dye loading are not well-established.

Purpose of the Study:

  • To characterize the impact of PF127 and DMSO concentrations on AM dye loading efficiency in slice cultures.
  • To determine optimal conditions for AM dye loading that maximize efficiency and preserve neuronal health.

Main Methods:

  • Investigated the effect of varying concentrations of PF127 and DMSO on AM dye loading in slice cultures.
  • Determined minimum effective concentrations for surfactant, solvent, and dye.
  • Utilized standard two-photon and confocal microscopy for monitoring neuronal activity.

Main Results:

  • Both PF127 and DMSO are essential for successful AM dye loading.
  • Lowering PF127 concentration significantly improved staining efficiency.
  • Reducing DMSO concentration to approximately 0.25% further enhanced loading efficiency.
  • Labeled neurons exhibited spontaneous and evoked calcium transients, with no observed deleterious effects on function for up to 24 hours.

Conclusions:

  • Optimized AM dye loading using lower PF127 concentrations enhances efficiency, preserves cell viability, and allows for long-term repetitive measurements.
  • Diluting the dye to a final concentration of 1μM reduces experimental costs.
  • This refined method is optimal for calcium imaging in slice cultures.