Polyclonal sensory neuron derivation from iPSCs as an efficient alternative to single clone strategies for pain relevant in vitro models
- 1Uniklinik RWTH Aachen
Abstract
Induced pluripotent stem cell (iPSC) workflows typically rely on monoclonal cultures, requiring multiple independent cell lines to compensate for clonal variability. This increases workload, cost, and limits scalability for translational applications. Here, we establish polyclonal reprogramming strategies using Sendai virus to generate clonally diverse iPSC cultures already during reprogramming through FACS, MACS, manual selection, or prolonged culture. Both monoclonal and FACS-derived polyclonal cultures reached pluripotency. Reprogramming transgenes were silenced, whereas Sendai virus (SeV) mRNA persisted across passages in both culture types; heat treatment at 38.5C markedly reduced SeV levels. Monoclonal and polyclonal cultures differentiated efficiently into neural crest-like cells and sensory neurons. As a second polyclonal strategy, monoclonal cultures were pooled at day5 of sensory neuron differentiation to generate percentage-controlled progenitor mixtures. TRPA1 protein expression has not been shown reliably in iPS-derived sensory neurons. We hypothesized that increased clonal diversity might facilitate detection of TRPA1 protein expression. Despite rigorous antibody validation, TRPA1 protein expression was detectable only diffusely across the whole cell area. Together, our results show that polyclonal iPSC strategies enable clonally inclusive generation of patient-specific sensory neurons. These workflows reduce early workload and provide a robust foundation for scalable drug-screening and patient-in-a-dish applications.
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BibTeX
@misc{neureiter2026polyclonal,
title = {{Polyclonal sensory neuron derivation from iPSCs as an efficient alternative to single clone strategies for pain relevant in vitro models}},
author = {A. Neureiter and L. Ruessmann and P. Hautvast and N. W. M. van den Braak and A. Stoppe and M. Haeusler and A. Lischka and K. Eggermann and A. Lampert},
howpublished = {bioRxiv},
year = {2026},
month = sep,
doi = {10.64898/2026.09.08.750017},
url = {https://www.biorxiv.org/content/10.64898/2026.09.08.750017v1},
note = {CC BY-NC},
}
RIS
TY - UNPB
TI - Polyclonal sensory neuron derivation from iPSCs as an efficient alternative to single clone strategies for pain relevant in vitro models
AU - A. Neureiter
AU - L. Ruessmann
AU - P. Hautvast
AU - N. W. M. van den Braak
AU - A. Stoppe
AU - M. Haeusler
AU - A. Lischka
AU - K. Eggermann
AU - A. Lampert
PY - 2026
DA - 2026/09/08
PB - bioRxiv
DO - 10.64898/2026.09.08.750017
UR - https://www.biorxiv.org/content/10.64898/2026.09.08.750017v1
AB - Induced pluripotent stem cell (iPSC) workflows typically rely on monoclonal cultures, requiring multiple independent cell lines to compensate for clonal variability. This increases workload, cost, and limits scalability for translational applications. Here, we establish polyclonal reprogramming strategies using Sendai virus to generate clonally diverse iPSC cultures already during reprogramming through FACS, MACS, manual selection, or prolonged culture. Both monoclonal and FACS-derived polyclonal cultures reached pluripotency. Reprogramming transgenes were silenced, whereas Sendai virus (SeV) mRNA persisted across passages in both culture types; heat treatment at 38.5C markedly reduced SeV levels. Monoclonal and polyclonal cultures differentiated efficiently into neural crest-like cells and sensory neurons. As a second polyclonal strategy, monoclonal cultures were pooled at day5 of sensory neuron differentiation to generate percentage-controlled progenitor mixtures. TRPA1 protein expression has not been shown reliably in iPS-derived sensory neurons. We hypothesized that increased clonal diversity might facilitate detection of TRPA1 protein expression. Despite rigorous antibody validation, TRPA1 protein expression was detectable only diffusely across the whole cell area. Together, our results show that polyclonal iPSC strategies enable clonally inclusive generation of patient-specific sensory neurons. These workflows reduce early workload and provide a robust foundation for scalable drug-screening and patient-in-a-dish applications.
ER -