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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: Buck-Wiese, Marie (VerfasserIn) , Liechocki, Sally (VerfasserIn) , Erfle, Holger (VerfasserIn) , Starkuviene-Erfle, Vytaute (VerfasserIn)
Dokumenttyp: Article (Journal)
Sprache:Englisch
Veröffentlicht: December 2025
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
Volltext
Verfasserangaben:Marie Buck-Wiese, Sally Liechocki, Holger Erfle, Vytaute Starkuviene
Beschreibung
Zusammenfassung: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.
Beschreibung:Online veröffentlicht: Dezember 2025
Gesehen am 03.02.2026
Beschreibung:Online Resource
ISSN:2472-5560
DOI:10.1016/j.slasd.2025.100283

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