Name: Bacterial Infection Quantitation Assay: A Method to Quantify Intraocular Bacterial Load in Mouse Model of Bacterial Endophthalmitis
Uploaded: 2023-04-30T14:51:57
Description: Source: Mursalin, M. H. et al., Intravitreal Injection and Quantitation of Infection Parameters in a Mouse Model of Bacterial Endophthalmitis. J. Vis. ...

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All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Harvesting tube preparation
<ol>
	<li>Place 1 mm sterile glass beads into 1.5 mL screw cap tubes.</li>
	<li>Sterilize these tubes in an autoclave on a dry setting. Let the tubes cool to room temperature before use.</li>
	<li>Add 10 mL of 1x sterile phosphate-buffered saline (PBS) to a sterile 15 mL centrifuge tube.</li>
	<li>Add 1 tablet of prote...

bacterial infection quantitation assay: a method to quantify intraocular bacterial load in mouse model of bacterial endophthalmitis

Source: Mursalin, M. H. et al., <a href="https://www.jove.com/v/61749">Intravitreal Injection and Quantitation of Infection Parameters in a Mouse Model of Bacterial Endophthalmitis.</a> J. Vis. Exp. (2021).
This video describes the technique of quantifying bacterial infection from eye homogenates obtained from a murine model of bacterial endophthalmitis caused by Bacillus cereus. This technique helps study the host immune response and its dependency on the host and ocular bacterial gene expression in murine models.

<img alt="Figure 1" src="/files/ftp_upload/20817/20817_Figure1.jpg" /> 
Figure 1: Harvesting mouse infected eye.&#160;Post-infection, after the desired time point, harvest infected mouse eyes using sterile tweezers. (A) PBS containing sterile glass beads. (B) Harvesting of the infected eye. (C) Harvest tube containing a mouse eye.
<img alt=...

Bacterial Infection Quantitation Assay: A Method to Quantify Intraocular Bacterial Load in Mouse Model of Bacterial Endophthalmitis

Source: Mursalin, M. H. et al., Intravitreal Injection and Quantitation of Infection Parameters in a Mouse Model of Bacterial Endophthalmitis. J. Vis. ...

Bacterial endophthalmitis is an inflammatory eye ailment that occurs when Bacillus cereus &#8212; a pathogenic bacteria &#8212; infects the intraocular region.
To quantify the bacterial infection, place a euthanized mouse model of bacterial endophthalmitis laterally on an operating table. Apply mild pressure under the eye to help detach the eyeball from the socket. Transfer the eyeball into a tube containing an appropriate buffer supplemented with protease inhibitor cocktail.
Homogenize the ocular tissue to disintegrate it, releasing bacteria in the suspension. The protease inhibitors present in the buffer inactivate the released cellular proteases to maintain bacterial viability. Next, serially dilute the bacterial suspension to obtain various concentrations of Bacillus cereus.
Take an angled culture plate containing the enriched agar medium and demarcate rows corresponding to each dilution concentration. Dispense a few drops from the bacterial suspension on the top of the designated row and allow it to flow down until it reaches the end of the plate, helping in the uniform spreading of bacteria. Flatten the plate and incubate.
During incubation, each bacterium uses the nutrients from agar to proliferate and form individual colonies. The number of colonies decreases, corresponding to an increase in the dilution of the bacterial suspension. Count the number of colonies in a particular row to quantify the ocular bacterial load.

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Bacterial Infection Quantitation Assay: A Method to Quantify Intraocular Bacterial Load in Mouse Model of Bacterial Endophthalmitis

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