{
  "abstract": "Transient receptor potential ankyrin 1 (TRPA1) is a polymodal sensory ion channel whose activity is influenced not only by chemical ligands but also by the physical properties of the plasma membrane. This raises the possibility that membrane-active compounds used routinely in cell biology may alter TRPA1 function. We investigated the acute effects of two widely used chemical transfection reagents, Lipofectamine 3000 and Mirus TransIT-293, on intracellular Ca2+; signaling and TRPA1 activity. For this, we monitored intracellular Ca2+; dynamics using ratiometric Fura-2 imaging in CHO cells stably expressing mouse TRPA1 (CHO-mTRPA1), parental CHO-WT cells, and primary mouse dorsal root ganglion (DRG) neurons. Transfection reagent preparations were applied at different concentrations under controlled temperature and low-flow conditions. The contribution of TRPA1 and Ca2+; influx was assessed using the selective TRPA1 inhibitor HC-030031 and the broad-spectrum Ca2+; channel blocker ruthenium red. We found that Lipofectamine 3000 induced concentration-dependent, irregular Ca2+; transients in CHO-mTRPA1 cells, while simultaneously inhibiting the constitutive TRPA1-dependent Ca2+; activity observed under basal conditions. Its inhibitory effect was evident at concentrations below those producing substantial cellular activation and was rapidly reversible after washout. At higher concentrations, Lipofectamine-induced Ca2+; responses were only partially suppressed by TRPA1 inhibition, indicating the involvement of additional mechanisms. Consistent with this, Lipofectamine also induced Ca2+; transients in CHO-WT cells and primary DRG neurons, where both extracellular Ca2+; influx and intracellular Ca2+; mobilization contributed to the responses. Analysis of the components of the Lipofectamine 3000 formulation further revealed distinct effects of Lipofectamine and the P3000 enhancer. Mirus TransIT-293 similarly induced Ca2+; transients in CHO-mTRPA1 cells and DRG neurons. In CHO-mTRPA1 cells, its response was concentration-dependent and strongly reduced by HC-030031, whereas the response in DRG neurons showed little sensitivity to TRPA1 inhibition. We conclude that chemical transfection reagents can acutely alter intracellular Ca2+; homeostasis and modulate TRPA1 activity. Their effects involve both TRPA1-dependent and TRPA1-independent mechanisms and differ substantially between formulations and cell types. These findings identify membrane-active transfection reagents as previously underappreciated modulators of sensory ion-channel function and highlight their potential to influence the interpretation of experiments performed in transfected cells.",
  "abstract_shown": true,
  "affiliations": [
    "KU Leuven"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 4,
  "authors": [
    {
      "name": "A. Milici"
    },
    {
      "name": "A. Segal"
    },
    {
      "name": "J. B. Startek"
    },
    {
      "affs": [
        0
      ],
      "name": "K. Talavera"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/5zxsqw2f5mndf.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.09.16.751488",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.09.16.751488v1.full",
  "id": "5zxsqw2f5mndf",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.09.16.751488v1",
  "language": null,
  "licence": "CC-BY-NC-ND",
  "licence_label": "CC BY-NC-ND",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by-nc-nd/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "cell biology"
    ],
    [
      "openalex",
      "2809|Ion Channels and Receptors"
    ]
  ],
  "openalex": "W7213595870",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.09.16.751488v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-09-16",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/5zxsqw2f5mndf.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:23:07Z",
      "native_id": "10.64898/2026.09.16.751488",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.09.16.751488v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:cell biology",
      "subfield": "cell-biology"
    }
  ],
  "title": "Chemical transfection reagents induce intracellular Ca2+ signals in TRPA1-expressing cells",
  "title_text": "Chemical transfection reagents induce intracellular Ca2+ signals in TRPA1-expressing cells",
  "type": "preprint",
  "updated_at": "2026-10-30T12:23:07Z",
  "url": "https://research.draconitic.com/paper/5zxsqw2f5mndf",
  "venue": "bioRxiv",
  "versions": []
}