High-resolution fluorescence measurements correlated to cellular traction forces

H. Hoorn, van, E.M. Spiesz, C. Storm, D. Noort, van, B. Ladoux, T. Schmidt

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Many cellular mechanosensory functions rely on molecular interactions related to cellular force exertion. Despite the rich biochemical knowledge in this field, the current mechanistic picture - that can be enhanced by high-resolution optical microscopy techniques - is still in its infancy. Here, we describe a simple yet universal approach to combine high-resolution optical microscopy techniques with traction force measurements. Etching 50 µm high spacers next to a micropillar array and inverting the array onto a thin coverslip before imaging enables the use of a short working distance objective. In our setup we use a spinning-disk confocal on an inverted microscope with a 100X, NA 1.4 objective. Our deflection detection resolution per micropillar is below 30 nm, corresponding to a force resolution of 500 pN. We validate our technique by imaging fixed and live mouse fibroblasts with dye-conjugated fibronectin micro-contact printed on the micropillars and immunostaining for paxillin and actin. We observe forces of up to 15 nN on individual pillars using a precise force-deflection relationship calibrated using scanning electron microscopy, tensile testing and finite element analysis. We quantified focal adhesion growth with increasing force of 4 ± 1 nN/µm2 and co-orientation of focal adhesion elongation and force direction within 2 ± 24 degrees. Furthermore, the cells remain viable during overnight live-cell imaging and stay adhered to the substrate. Our novel approach opens up the combined application of traction force measurements and super-resolution optical microscopy techniques.
Original languageEnglish
Pages (from-to)193a-
Number of pages1
JournalBiophysical Journal
Volume104
Issue number2
DOIs
Publication statusPublished - 2013

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