a zeiss axio observer z1  (Carl Zeiss)

Journal: Nucleic Acids Research

Article Title: RecN and RecA orchestrate an ordered DNA supercompaction response following ciprofloxacin-induced DNA damage in Escherichia coli

doi: 10.1093/nar/gkaf437

Figure Lengend Snippet: DNA supercompaction in wild-type E. coli cells (KV21) after CIP exposure. All cells were grown in LB at 37°C and imaged at 2-min intervals using live-cell imaging. ( A and B ) Cells were immobilized in microfluidic channel slides and imaged using widefield microscopy. (A) Representative images of wild-type cells with HU-mCherry fluorescence (green) at 0, 8, and 16 min after CIP exposure, showing DNA distribution; scale bar: 5 μm. (B) Analysis of DNA distribution along the cells’ long axis before and after CIP, quantified by measuring the distance between the outer bounds of symmetrical fluorescence peaks at 80% of maximum averaged intensity for each time point. Results are averaged from 34 to 141 cells from a single representative biological replicate (see “Materials and methods” section for detailed explanation). ( C and D ) Cells were immobilized on agar pads and imaged using spinning disk microscopy. Results shown are from images captured 12, 14, 16, and 18 min after CIP exposure, averaged from 40 tracked cells from a single representative biological replicate. (C) Average HU-mCherry fluorescence intensity along the cells’ long axis. (D) Kymograph heat map of relative HU-mCherry intensity distribution over time. ( E ) DNA compaction phenotype distribution for baseline unchallenged cells (dots) and cells after CIP exposure (lines). Dots and lines represent means from three biological replicates, while error bars and shaded regions indicate standard deviation. A.u., arbitrary unit.

Article Snippet: To examine the cells’ immediate response to CIP exposure, we employed a microfluidic setup to image cells using a Zeiss Axio Observer Z1 widefield inverted microscope with a 63x oil objective (Zeiss Plan Apochromat 1.4 NA, DIC), a Colibri 7 LED light source, a Hamamatsu ORCA-Flash4.0 V3 digital CMOS camera, as well as a heated incubation chamber and mounting frame, both maintained at 37°C.

Techniques: Live Cell Imaging, Microscopy, Fluorescence, Standard Deviation

Journal: Nucleic Acids Research

Article Title: RecN and RecA orchestrate an ordered DNA supercompaction response following ciprofloxacin-induced DNA damage in Escherichia coli

doi: 10.1093/nar/gkaf437

Figure Lengend Snippet: ΔrecN cells exhibit limited nucleoid reorganization after CIP exposure. All cells were grown in LB at 37°C and imaged at 2-minute intervals using live-cell imaging. ( A and B ) Cells (MR16) were immobilized in microfluidic channel slides and imaged using widefield microscopy. (A) Representative images of ΔrecN cells at 0, 8, and 16 min after CIP exposure, showing HU-mCherry fluorescence (green) to represent DNA distribution. Scale bar: 5 μm. ( B ) Analysis of DNA distribution along the cells’ long axis before and after CIP, quantified by measuring the distance between the outer bounds of symmetrical fluorescence peaks at 80% of maximum averaged intensity for each time point. Results are averaged from 17 to 61 cells from a single representative biological replicate (see “Materials and methods” section for detailed explanation). ( C and D ) Cells (KV68) immobilized on agar pads and imaged using spinning disk microscopy. Results are shown from images captured 12, 14, 16, and 18 min after CIP exposure, averaged from 40 tracked cells from a single representative biological replicate. (C) Average HU-mCherry fluorescence intensity along the cells’ long axis. (D) Kymograph heat map of relative HU-mCherry intensity distribution over time. ( E ) DNA compaction phenotype distribution for ΔrecN cells (KV68) at unchallenged baseline (dots) and after CIP exposure (lines). Dots and lines represent means from three biological replicates; error bars and shaded regions indicate standard deviation. A.u., arbitrary unit.

Article Snippet: To examine the cells’ immediate response to CIP exposure, we employed a microfluidic setup to image cells using a Zeiss Axio Observer Z1 widefield inverted microscope with a 63x oil objective (Zeiss Plan Apochromat 1.4 NA, DIC), a Colibri 7 LED light source, a Hamamatsu ORCA-Flash4.0 V3 digital CMOS camera, as well as a heated incubation chamber and mounting frame, both maintained at 37°C.

Techniques: Live Cell Imaging, Microscopy, Fluorescence, Standard Deviation