Comparative analysis of antibody-mediated loss-of-function versus gene knock-out and knock-down
In this study we compare three methods for manipulating cell function: RNA interference (RNAi), CRISPR-Cas9 gene knock-out, and antibody-mediated loss-of-function. We have focused on analyzing changes in cell-matrix adhesion via targeting two key regulators, Talin1 (TLN1) and Kindlin-2 (KD2). Adhesi...
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| Hauptverfasser: | , , , |
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| Dokumenttyp: | Article (Journal) |
| Sprache: | Englisch |
| Veröffentlicht: |
December 2025
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| In: |
SLAS discovery
Year: 2025, Jahrgang: 37, Pages: 1-11 |
| ISSN: | 2472-5560 |
| DOI: | 10.1016/j.slasd.2025.100283 |
| Online-Zugang: | Verlag, kostenfrei, Volltext: https://doi.org/10.1016/j.slasd.2025.100283 Verlag, kostenfrei, Volltext: https://www.sciencedirect.com/science/article/pii/S2472555225000760 
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| Verfasserangaben: | Marie Buck-Wiese, Sally Liechocki, Holger Erfle, Vytaute Starkuviene |
MARC
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| 520 | |a In this study we compare three methods for manipulating cell function: RNA interference (RNAi), CRISPR-Cas9 gene knock-out, and antibody-mediated loss-of-function. We have focused on analyzing changes in cell-matrix adhesion via targeting two key regulators, Talin1 (TLN1) and Kindlin-2 (KD2). Adhesion-relevant phenotypic assays revealed distinct temporal onset dynamics for each method. RNAi and CRISPR-Cas9 effectively reduced target mRNA and protein levels. In contrast, antibody transfection induced phenotypic changes without altering target expression, suggesting direct intracellular antibody-target interaction. Transcriptome analysis demonstrated that antibody transfection and CRISPR-Cas9 induced fewer deregulated mRNAs than RNAi. Furthermore, transfected antibodies and sgRNAs shared 30 % and 70 % of deregulated transcripts to their negative controls, respectively. Whereas only 10 % of overlap was recorded between targeting and control siRNAs. Our findings emphasize the importance of considering method-specific temporal dynamics of on-target phenotype appearance and off-target manifestation. Additionally, they highlight intracellular delivered antibodies as a valuable alternative for validating and complementing genetic approaches. | ||
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