Live-Cell Time-Lapse Imaging of Bdellovibrio bacteriovorus Predation on E. coli

Samenvatting

Source: Makowski, Ł., et al. Live-Cell Imaging of the Life Cycle of Bacterial Predator Bdellovibrio bacteriovorus using Time-Lapse Fluorescence Microscopy. J. Vis. Exp. (2020)

This video demonstrates time-lapse fluorescence imaging of Bdellovibrio bacteriovorus preying on immobilized E. coli beneath an agarose pad, enabling real-time visualization of intracellular replication and progeny release for studying bacterial predation and host-pathogen interactions.

Protocol

1. Conduction of time-lapse fluorescence microscopy

1. Place the 35 mm dish in the Petri dish holder such that it will not move during the course of the experiment. Place one drop of immersion oil (1.518 refractive index) onto the objective as well as the bottom of the 35 mm dish.
2. Mount the holder onto the stage of the inverted microscope in the microscope chamber.
3. Set up the options in the microscope control software to collect multiple images at multiple stage positions over time.
   1. Set the magnification (100x) and polychromic lens (GFP (green fluorescent protein)/mCherry). Using an eyepiece and brightfield (....

Materialen

Lijst van materialen gebruikt in dit artikel

Naam	Bedrijf	Catalogusnummer	Opmerkingen
CertifiedMolecular Biology Agarose	BIO-RAD	161-3100	Low fluorescence agarose for agarose pad
Fiji	ImageJ	https://imagej.net/Fiji	Open source image processing package
Glass Bottom Dish 35 mm	ibidi	81218-200	Uncoated glass
Microscope	GE	DeltaVision Elite	Microtiter Stage, ultimate focus laser module, DV Elite CoolSnap HQ2 Camera, SSI assembly FP DV, kit obj. Oly 100x oil 1.4 NA, prism Nomarski 100x LWD DIC, ENV ctrl IX71 uTiter opaQ 240 V
Start SoftWoRx	GE		Manufacturer-supplied imaging software