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Chemical Force Spectroscopy on single living cells

Chemical Force Spectroscopy on single living cells. E. Dague, D. Alsteens, J.P. Latgé, Y. Dufrêne. Aspergillus hyphae and conidia as seen by SEM. Kaminsky et al. , 2006. Mycological Research. Classical way of probing microbial hydrophobicity.

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Chemical Force Spectroscopy on single living cells

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  1. Chemical Force Spectroscopy on single living cells E. Dague, D. Alsteens, J.P. Latgé, Y. Dufrêne

  2. Aspergillus hyphae and conidia as seen by SEM Kaminsky et al., 2006. Mycological Research

  3. Classicalway of probingmicrobial hydrophobicity Not specific of hydrophobic character and under electrostatic interactions influence (Mozes, Rouxhet 1987 ; van Loosdrecht et al., 1990 ;Gaboriaud et al., 2006) • Hydrophobicity • MATH, MATS • HIC • Adhesion test • Contact angle vortex 1 min

  4. Probing hydrophobic forces at the nanoscale Increasing CH3 surface fraction

  5. 10 10 8 8 6 6 Adhesion force (nN) 4 4 2 2 0 0 0 0 20 20 40 40 60 60 80 80 100 100 Surface fraction CH (%) (%) 3 3 Hydrophobicity quantification and calibration 100 % CH 3 75 % CH 3 30 50 % CH 3 25 % CH 3 0 % CH 3 20 Frequency (%) 10 0 0 2 4 6 8 10 Adhesion force (nN) Hydrophobicity calibration Alsteens, Dague et al., Langmuir 2007

  6. water single cell tape membrane sampleholder Single Aspergillus conidia trapped in a pore as seen by contact mode AFM 500 nm 500 nm

  7. Hydrophobic Force Microscopy of A. fumigatus 5 1000 pN 4 100 nm Dal : native 3 Frequency (%) 2 1 0 100 nm 100 nm 0 1000 2000 3000 4000 5000 6000 Adhesion force (pN)

  8. Hydrophobic Force Microscopy of A. fumigatus 5 1000 pN 4 100 nm 3 Dal : native 2 1 100 nm 100 nm 0 Frequency (%) 15 1000 pN 10 100 nm 5 Dal + NaOH 1M 0 100 nm 100 nm 0 1000 2000 3000 4000 5000 6000 Adhesion force (pN)

  9. Hydrophobic Force Microscopy of A. fumigatus 5 1000 pN 4 100 nm Dal : native 3 2 1 100 nm 100 nm 0 15 1000 pN 10 100 nm Frequency (%) 5 Dal + NaOH 1M 100 nm 100 nm 0 15 10 1000 pN Mutant without hydrophobins 100 nm 5 0 100 nm 100 nm 0 1000 2000 3000 4000 5000 6000 Adhesion force (pN) Dague, Alsteens et al., Nanoletters 2007

  10. 100 nm

  11. Adhesion force map revealing highly correlated structural and hydrophobic heterogeneities 100 nm 100 nm Dague, Alsteens et al., Nanoletters 2007

  12. Aspergillus hyphae and conidia as seen by SEM Kaminsky et al., 2006. Mycological Research

  13. Real time imaging of conidia during germination Germination Dague et al., Biophysical Journal 2007

  14. Structural dynamics of single germinating conidia 200 nm 200 nm 200 nm 20 min 60 min 0 min 200 nm 200 nm 200 nm 120 min 120 min 180 min

  15. Evolution of hydrophobicity with germination time 100 nm 100 nm 100 nm 60 min 90 min 0 min 100 nm 200nm 120 min 200 nm 210 min 180 min Dague et al., Biophysical Journal 2007

  16. Take Home Message • Hydrophobicity can be probed and quantified at the nanoscale • Direct visualization of correlated morphological and chemical evolution • Biological process (germination), chemical modifications can be observed, in real time

  17. D. Alsteens J.P. Latgé • Y. Dufrêne

  18. Chemical Force Spectroscopy on single living cells E. Dague, D. Alsteens, J.P. Latgé, Y. Dufrêne

  19. Toward a global bio-surface analysis :Combined use of AFM, XPS and SIMS 0.02 A B 2.5.10-2 Intensity (a. u.) XPS N peak Rod B CN- NI, SIMS 2.10-2 Dal Normalized intensities i/itot 1.5.10-2 0.01 Rod A Rod AB %CPr, XPS C D 1.10-2 C2H3O2- NI, SIMS 15 20 25 30 35 1.5.10-3 0.00 WT ∆RodA ∆RodA ∆RodB ∆RodB 1.10-3 Dague et al., Langmuir 2008 %CPs, XPS 5.10-4 10 15 20 25 30

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