Overview
Chitozen is a chitosan-coated coverslip compatible with 6-channel sticky slides.
It is very helpful if you want to
- image bacteria both still and alive under the microscope
- Maintain bacteria in a same focal plane for imaging while preserving their physiology
-
Renew culture medium and change the growth condition (e.g. antibiotics, chemicals, inhibitors) during the experiment and directly observe, in real-time, the bacteria new comportment under the microscope.
A technology designed by
Tâ
m
Mignot
,
Olivier Theodoly
,
Amandine Desorme
,
Guillaume Sudre
and
Laurent David
.
Published in mBio / ScienceAdvances
E. coli but non only
Chitozen is efficient on the following bacteria:
- E. coli
- Bacillus subtilis
-
Caulobacter crescentus
- Corynebacterium glutamicum
- Helicobacter pylori
- Mycobacterium smegmatis
- Myxococcus xanthus
- Pseudomonas aeruginosa
- Pseudomonas fluorescens
- Salmonella
- Staphylococcus aureus
- Vibrio cholerae
We update this list regularly according to feedback provided by researchers who use Chitozen.
Videos & images
© 2019 Tréguier et al. This content is distributed under the terms of the Creative Commons Attribution 4.0 International license.
Visualization of Pal mCherry at septum in
E. coli
(W3110 Pal mCherry), by 3D SIM microscopy, in M9 medium and using Chitozen.
Credit : Amandine Desorme - LCB - CNRS - 2021
Features
High adherence
Maintains bacteria in a similar focal plane for imaging
Preserves cell physiology
The chitosan coating is free from bacterio-static effects and does not affect bacteria geometry
Perfusable
Change the medium or add compounds throughout experiment to assess real-time changes in bacterial behavior
Easy-to-use
Suitable as an easy starting tool to get trained on bacterial imaging
Time-saving
Test up to 6 parallel conditions on one slide
Protocol
ASSEMBLE COVERSLIP & STICKY-SLIDE
LOAD 1 SAMPLE PER CHANNEL
CENTRIFUGE OR LEAVE BACTERIA TO SETTLE
IMAGE
Going further: perfusing the system with flow
Feedback from users
Chitozen has been successfully tested by users on the following bacteria:
E. coli
> Caulobacter crescentus
> Corynebacterium glutamicum
Helicobacter pylori
> Mycobacterium smegmatis
Bacillus subtilis
Pseudomonas fluorescens
> Salmonella
> Staphylococcus aureus
These results supplement those obtained by Tâm Mignot and al. with:
E. coli
Vibrio cholerae
Myxococcus xanthus
Pseudomonas aeruginosa
Feedback by Emily Helgesen
Oslo University Hospital, 2022
Live cell imaging of E. coli in response to drug addition on Chitozen slides
AB1157 E. coli expressing DNA marker HU-mCherry were imaged at 37oC with perfusion of ½ LB at a flow rate of 2 ml/min. Drug X was added at time point 35 minutes and removed at time point 60 minutes. Credit: Emily Helgesen - 2022
Live imaging of E. coli expressing a protein associated with DNA replication on Chitozen slides
AB1157 E. coli cells expressing SeqA-YFP (pseudo colored green), a protein associated with DNA replication, were imaged at 37oC over 60 minutes without perfusion of medium.
Credit: Emily Helgesen – 2022
Feedback by Bianca Sclavi
Sorbonne University, 2021
Effects of cell division inhibitor cephalexin on E. coli growth cultured on Chitozen slides
Credit: Bianca Sclavi - 2021
Feedback by Itzhak Fishov
Ben-Gurion University of the Negev, 2022
E. coli spheroblasts bound to Chitozen 
E. coli MG1655 with chromosomally encoded HU-eGFP under the native promoter were imaged with perfusion at a flow rate of 0.05 mL/min. Credit: Itzhak Fishov – 2022
Feedback by Maxence Vincent
University of Oxford, Jan. 2022
Tracking of DNA repair proteins in live cells of Escherichia coli
"Cells expressing HaloTagged DNA repair proteins were loaded onto the chitosan-coated coverslip and imaged with TIRF microscopy. The Chitozen technology enables tracking single-molecules while changing environmental conditions (e.g: M9 or M9 + mutagen). Computation of the diffusion coefficients provides insights into how proteins change their mobility upon DNA damage".
Feedback by Jerome Rech
LMGM-CBI-CNRS, Toulouse, 2022
Time-lapse images of E. coli growing: Localization of F-plasmid
Cells are observed in phase contrast (top), in the blue and yellow channels for fluorescence microcopy to observe phase contrast, nucleoid (DAPI) or F-plasmid (mVenus tag), respectively.
Overlays of phase contrast and blue or yellow channel are shown on middle or bottom panels, respectively.
Cells were grown at 30°C in supplemented M9 minimal media containing Cystein and DAPI under continuous flow. Scale bar (2 µm). Credit: Jerome Rech - 2022
Feedback by Andrea Vettiger
Harvard University, June 202
"I used the Chitozen coverslips together with Corynebacterium glutamicum and Staphylococcus aureus. We appreciated the good adherence of the bacteria on the surface and the versatility of the tool."
Technical information
Assay compatibility:
> Fluorescence
> Co-cultures (bacterial predators, immune cells)
Addition of external factors (e.g. antibiotics, chemicals, inhibitors)
> Static or dynamic conditions
Imaging modes: phase-contrast, epifluorescence, confocal, super-resolution microscopy, atomic force microscopy (AFM)*
*documentation & example pictures available on request
Experimental outputs:
> Behavioural changes: growth, elongation, cell division, fitness, colony/biofilm formation, etc
> Single molecule imaging
Kit description
5 standard (25 x 75 mm) chitosan-coated coverslips with 5 sticky slides.
Each Chitozen coverslip allows up to 6 assays using bottomless 6-channel sticky slides.
Lifetime: up to 12-month storage at room temperature, shielded from direct sunlight.
Resources:
>
FAQ
>
Technical Datasheet
>
Safety Datasheet
Story
Tâm Mignot, Olivier Théodoly, Amandine Desorme, Guillaume Sudre, Laurent David
Esther Graudens
New projects at Idylle
And if you are interested in seeing what you can do with chitosan-coated coverslips, you can have a look at this study published by the team of developers in 2019. They've managed to use chitosan-coated coverslips to promote the growth of cells without any deleterious effect on their physiology, allowing them to measure the antibiotic susceptibility of a diversity of clinical strains with an excellent accuracy in a very short period of time. Read all about it here: mBio, 2019"