Ex vivo Pig Lung Model of Biofilm: A Technique to Develop Lung Model to Study Bacterial Biofilms on Pig Bronchiolar Tissue

Abstract

Source: Harrington, N.E. et al., Antibiotic Efficacy Testing in an Ex vivo Model of Pseudomonas aeruginosa and Staphylococcus aureus Biofilms in the Cystic Fibrosis Lung. J. Vis. Exp. (2021). 

This video describes the protocol for developing an ex vivo pig lung model of bacterial biofilm on infected bronchiolar tissue. Biofilms are bacterial aggregates embedded in a matrix adherent to a surface. This model is used to study the antibiotic susceptibility of bacteria at different stages of biofilm formation.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Dissection and infection of ex vivo pig lung (EVPL) tissue

1. Make SCFM for use with EVPL tissue, follow the recipe for 1 L modified SCFM supplied in Table 1.
2. Prior to dissection, prepare an agar plate/s of required bacterial strain/s for infection using whatever agar is standard in the lab for P. aeruginosa/S. aureus (e.g., lysogeny broth + 1.2% agar).
3. Calculate how many porcine bronchiolar tissue pieces are required for the experi....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Insulin syringes - 0.5 mL with 29G needle attached.	VWR	BDAM324892
24-well culture plates
70% ethanol or similar for surface sterilization and flaming of dissection equipment
Agar plates to prepare streaks of P. aeruginosa/S. aureus (any suitable medium)
Agarose
Aluminum foil - pre-sterilised by autoclaving - to cover the chopping board on whcih you wil dissect lungs.
Breathe-easy or Breathe-easier sealing membrane for multiwell plates	Diversified Biotech	BEM-1 or BERM-2000
Bunsen burner
Chopping board - we recommend a plastic board to allow for easy decontamination with alcohol.
Coolbox to transport lungs to lab
Dissection scissors in different sizes
Dulbecco's modified Eagle medium (DMEM)
Large pallet knife
Mounted razor blades
Petri dishes
Plastic chopping board and aluminium foil to create a sterile and cleanable dissection surface
Roswell Park Memorial Institute (RPMI) 1640 medium
SCFM			Ingredients as listed in Table 1
Selection of forceps (blunt tips recommended)
Suitable containers for disposing of contaminated sharps and pig lung tissue, according to your institution's health & safety policies.