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Calcium ions (Ca2+) serve as crucial secondary messengers in numerous signaling cascades in plants, similar to their role in animals. One of the earliest responses in the activation of immune responses is a rapid, transient increase in cytosolic Ca2+ ([Ca2+]cyt). Measuring the spatial and temporal changes in [Ca2+]cyt is essential for understanding the details of immune responses to various pathogens and pests. We present a straightforward assay for measuring [Ca2+]cyt using the genetically encoded Ca2+ indicator, GCaMP, and a fluorescence microplate reader. This assay allows for the assessment of changes in [Ca2+]cyt with high temporal resolution following various elicitor treatments. Additionally, we describe a method to analyze [Ca2+]cyt using a dexamethasone-inducible effector expression system (proDEX::AvrRpm1). Due to the simplicity of these methods, they are also suitable for high-throughput screening.

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Plate Reader-Based Quantification of Cellular Calcium Dynamics in Immune Responses Using the Genetically Encoded Calcium Indicator GCaMP

  • 实验方案

  • 发表日期：

    2026年

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  • Hyunsuh Lee,
  • Robin Hon Ming Goh,
  • Wolfgang Moeder,
  • Keiko Yoshioka

摘要

Calcium ions (Ca2+) serve as crucial secondary messengers in numerous signaling cascades in plants, similar to their role in animals. One of the earliest responses in the activation of immune responses is a rapid, transient increase in cytosolic Ca2+ ([Ca2+]cyt). Measuring the spatial and temporal changes in [Ca2+]cyt is essential for understanding the details of immune responses to various pathogens and pests. We present a straightforward assay for measuring [Ca2+]cyt using the genetically encoded Ca2+ indicator, GCaMP, and a fluorescence microplate reader. This assay allows for the assessment of changes in [Ca2+]cyt with high temporal resolution following various elicitor treatments. Additionally, we describe a method to analyze [Ca2+]cyt using a dexamethasone-inducible effector expression system (proDEX::AvrRpm1). Due to the simplicity of these methods, they are also suitable for high-throughput screening.