Assembly of live micro-organisms on microstructured PDMS stamps by convective/capillary deposition for AFM bio-experiments

Dague, Étienne; Jauvert, Eric; Laplatine, Loïc; Viallet, B.; Thibault, Christophe; Ressier, Laurence

doi:10.1088/0957-4484/22/39/395102

Cited by 60 publications

(48 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These microchambers can easily be loaded by convective/capillary deposition of yeast cells. This versatile method was validated by AFM nanomechanical measurements of yeast cells and germinated Aspergillus conidia in growth media (27). In addition, we investigated the role of the transcription factor Msn2/Msn4 in this AFM analysis of ethanol stress.…”

mentioning

confidence: 99%

Evidence for a Role for the Plasma Membrane in the Nanomechanical Properties of the Cell Wall as Revealed by an Atomic Force Microscopy Study of the Response of Saccharomyces cerevisiae to Ethanol Stress

Schiavone

Formosa-Dague

Elsztein

et al. 2016

Appl Environ Microbiol

View full text Add to dashboard Cite

A wealth of biochemical and molecular data have been reported regarding ethanol toxicity in the yeast Saccharomyces cerevisiae. However, direct physical data on the effects of ethanol stress on yeast cells are almost nonexistent. This lack of information can now be addressed by using atomic force microscopy (AFM) technology. In this report, we show that the stiffness of glucosegrown yeast cells challenged with 9% (vol/vol) ethanol for 5 h was dramatically reduced, as shown by a 5-fold drop of Young's modulus. Quite unexpectedly, a mutant deficient in the Msn2/Msn4 transcription factor, which is known to mediate the ethanol stress response, exhibited a low level of stiffness similar to that of ethanol-treated wild-type cells. Reciprocally, the stiffness of yeast cells overexpressing MSN2 was about 35% higher than that of the wild type but was nevertheless reduced 3-to 4-fold upon exposure to ethanol. Based on these and other data presented herein, we postulated that the effect of ethanol on cell stiffness may not be mediated through Msn2/Msn4, even though this transcription factor appears to be a determinant in the nanomechanical properties of the cell wall. On the other hand, we found that as with ethanol, the treatment of yeast with the antifungal amphotericin B caused a significant reduction of cell wall stiffness. Since both this drug and ethanol are known to alter, albeit by different means, the fluidity and structure of the plasma membrane, these data led to the proposition that the cell membrane contributes to the biophysical properties of yeast cells. IMPORTANCEEthanol is the main product of yeast fermentation but is also a toxic compound for this process. Understanding the mechanism of this toxicity is of great importance for industrial applications. While most research has focused on genomic studies of ethanol tolerance, we investigated the effects of ethanol at the biophysical level and found that ethanol causes a strong reduction of the cell wall rigidity (or stiffness). We ascribed this effect to the action of ethanol perturbing the cell membrane integrity and hence proposed that the cell membrane contributes to the cell wall nanomechanical properties. T he yeast Saccharomyces cerevisiae is a remarkable ethanol producer that is also very sensitive to its main fermentative product. At low to moderate concentrations (5 to 7%), ethanol mainly affects the growth rate, and at higher concentrations (Ͼ10%), it can strongly impair cell integrity, eventually leading to cell death with features of apoptosis (1). These inhibitory and toxic effects are ascribed to the fact that ethanol alters cell membrane fluidity and dissipates the transmembrane electrochemical potential, thereby creating permeability to ionic species and causing leakage of metabolites (2). Recent works using lipidomic methodologies confirmed the relationship between the composition of lipids, notably ergosterol and unsaturated fatty acids, and ethanol tolerance (3, 4). Moreover, as it diffuses freely into cells, ethanol at high concen...

show abstract

mentioning

confidence: 99%

Evidence for a Role for the Plasma Membrane in the Nanomechanical Properties of the Cell Wall as Revealed by an Atomic Force Microscopy Study of the Response of Saccharomyces cerevisiae to Ethanol Stress

Schiavone

Formosa-Dague

Elsztein

et al. 2016

Appl Environ Microbiol

View full text Add to dashboard Cite

show abstract

“…This is because that as the scan size decreased, the influence of cell membrane movement decreased and this cause the image quality increased. Dague et al [28] used PDMS stamps to trap microbial cells (Saccharomyces cerevisiae yeasts and Aspergillus fumigatus fungal spores). Microbial cells have thicker, stiffer cell walls, which allow easier acquisition of good quality of AFM images.…”

Section: Resultsmentioning

confidence: 99%

Atomic force microscopy imaging of live mammalian cells

Liu

et al. 2013

Sci. China Life Sci.

View full text Add to dashboard Cite

Atomic force microscopy (AFM) was used to examine the morphology of live mammalian adherent and suspended cells. Time-lapse AFM was used to record the locomotion dynamics of MCF-7 and Neuro-2a cells. When a MCF-7 cell retracted, many small sawtooth-like filopodia formed and reorganized, and the thickness of cellular lamellipodium increased as the retraction progressed. In elongated Neuro-2a cells, the cytoskeleton reorganized from an irregular to a parallel, linear morphology. Suspended mammalian cells were immobilized by method combining polydimethylsiloxane-fabricated wells with poly-L-lysine electrostatic adsorption. In this way, the morphology of a single live lymphoma cell was imaged by AFM. The experimental results can improve our understanding of cell locomotion and may lead to improved immobilization strategies.atomic force microscopy, cell locomotion, lamellipodia, cytoskeleton, polydimethylsiloxane Citation:Li M, Liu L Q, Xi N, et al. Atomic force microscopy imaging of live mammalian cells.

show abstract

“…The drawback of these lateral forces is that the biological sample has to be firmly immobilized on the surface to overcome them (figure 1). Thus strategies were developed to perform non denaturing immobilization [15][16][17][18] . It must be noticed that mammalian cells are usually spreading on their support and that no special immobilization procedure is needed ( figure 1).…”

Section: Imagingmentioning

confidence: 99%

Atomic Force Microscopy to Explore Electroporation Effects on Cells

Dague

2016

Handbook of Electroporation

View full text Add to dashboard Cite

Atomic Force Microscopy (AFM) is more and more used in life science. Its ability to provide images of living cells as well as mechanical or adhesion maps makes it a technology that cannot be ignored. In the context of electroporation (EP) which undoubtly affects the cells membrane or wall, a technology able to probe the cell surface is more than interesting. This chapter describes the principle of the AFM technology and especially the latest multiparametric imaging modes developed recently. It then demonstrates that AFM can be used to probe cell's morphology modifications induced by electric pulses. We then show that EP modifies cell's nanomechanical properties and that the actin cytoskeleton plays a major role in this process. Finally we shed light on the effects of EP on bacteria as probed by AFM.In this latest example it must be noticed that no mechanical modifications are induced, but the adhesion properties of the bacteria are dramatically reduced by Pulsed Eelectric Fields (PEF).Altogether the chapter shows the interest of applying AFM on cells exposed to EP, in order to get a better fundamental understanding of EP effect on cells or bacteria.

show abstract

Assembly of live micro-organisms on microstructured PDMS stamps by convective/capillary deposition for AFM bio-experiments

Cited by 60 publications

References 40 publications

Evidence for a Role for the Plasma Membrane in the Nanomechanical Properties of the Cell Wall as Revealed by an Atomic Force Microscopy Study of the Response of Saccharomyces cerevisiae to Ethanol Stress

Evidence for a Role for the Plasma Membrane in the Nanomechanical Properties of the Cell Wall as Revealed by an Atomic Force Microscopy Study of the Response of Saccharomyces cerevisiae to Ethanol Stress

Atomic force microscopy imaging of live mammalian cells

Atomic Force Microscopy to Explore Electroporation Effects on Cells

Contact Info

Product

Resources

About