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Developing a Robust Multiround HCR-FISH Method Modified for Caenorhabditis elegans.

Xiaoxiong Yang1, Jingxuan Zeng2, Chung-Kuan Chen2

  • 1Department of Computational Biology and Medical Sciences, Graduate School of Frontier Sciences, University of Tokyo, Chiba, Japan.

Genes to Cells : Devoted to Molecular & Cellular Mechanisms
|March 10, 2026
PubMed
Summary
This summary is machine-generated.

Researchers developed an iterative FISH method for whole-body C. elegans, improving sample retention to over 99% and enabling multiplexed gene profiling alongside neural activity measurements.

Keywords:
C. elegansHCRsequential FISHwhole‐brain imaging

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Area of Science:

  • Neuroscience
  • Molecular Biology
  • Genetics

Background:

  • Multiplexed gene expression visualization at cellular resolution is key for understanding biological systems.
  • Sequential fluorescent in situ hybridization (FISH) is used for gene profiling in complex tissues.
  • Caenorhabditis elegans is a vital model organism in neuroscience and genetics, but lacks optimized iterative FISH protocols.

Purpose of the Study:

  • To develop and optimize a multiround hybridization chain reaction FISH method for whole-body C. elegans.
  • To improve sample retention during iterative probing cycles.
  • To enable correlation of multiplexed molecular data with functional measurements in the same individual.

Main Methods:

  • Developed a multiround hybridization chain reaction FISH protocol tailored for C. elegans.
  • Chemically attached hydrogel-embedded samples to coverglass to enhance retention.
  • Validated the protocol's efficiency and reproducibility through multiple probing cycles.

Main Results:

  • Achieved over 99% sample retention across multiple probing cycles after implementing coverglass attachment.
  • Maintained high signal fidelity with over 86% detection efficiency and 80% spot-matching reproducibility.
  • Successfully applied the protocol to profile multiple genes in a single C. elegans individual post-neural activity measurement.

Conclusions:

  • The optimized iterative FISH protocol significantly enhances sample retention and data fidelity in C. elegans.
  • This accessible method provides a powerful framework for correlating gene expression with functional data at cellular resolution.
  • Facilitates advanced molecular and functional studies in C. elegans neuroscience and genetics.