Home › Microbiology › Isolation of Single Intracellular Bacterial Communities Generated from a Murine Model of Urinary Tract Infection for Downstream Single-cell Analysis
Microbiology JoVE (Open Access) Citable · DOI

Isolation of Single Intracellular Bacterial Communities Generated from a Murine Model of Urinary Tract Infection for Downstream Single-cell Analysis

DOI: 10.3791/58829-v
What you'll learn
  • ✓ Prepare glass capillaries for single-cell isolation from infected tissue
  • ✓ Harvest bladder epithelial cells from murine UTI model
  • ✓ Isolate pure intracellular bacterial communities for downstream analysis
  • ✓ Validate IBC purity and viability post-isolation
Protocol

This protocol describes a simple method of isolating single, infected-bladder epithelial cells from a murine model of urinary tract infection.

Difficulty
advanced
Total time
~2–3 hours per mouse (including infection model establishment not shown)
Model organism
Mouse (murine UTI model)
Biosafety
BSL-2

Steps

1
Prepare glass capillaries for cell isolation

Draw and shape glass capillaries to appropriate diameter for single infected epithelial cell capture. This preparation enables precise micromanipulation during downstream IBC isolation.

▶ 00:41
2
Harvest bladder epithelial cells from infected tissue

Dissect infected bladder tissue and isolate individual epithelial cells using enzymatic digestion or mechanical dissociation. Prepare cell suspension for single-cell selection.

▶ 02:06
3
Isolate intracellular bacterial communities from single cells

Use prepared glass capillaries and micromanipulation to extract single infected cells and release their intracellular bacterial communities. Ensure separation of IBCs from host cell debris.

▶ 03:53
4
Validate IBC purity and suitability for analysis

Assess harvested intracellular bacterial communities for purity, contamination, and viability. Confirm readiness for single-cell genomics, transcriptomics, or other downstream applications.

▶ 05:18
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