Investigating IL-1β Secretion Using Real-Time Single-Cell Imaging

Methods Mol Biol. 2016;1417:75-88. doi: 10.1007/978-1-4939-3566-6_4.

Abstract

The pro-inflammatory cytokine interleukin (IL)-1β is an important mediator of the inflammatory response. In order to perform its role in the inflammatory cascade, IL-1β must be secreted from the cell, yet it lacks a signal peptide that is required for conventional secretion, and the exact mechanism of release remains undefined. Conventional biochemical methods have limited the investigation into the processes involved in IL-1β secretion to population dynamics, yet heterogeneity between cells has been observed at a single-cell level. Here, greater sensitivity is achieved with the use of a newly developed vector that codes for a fluorescently labelled version of IL-1β. Combining this with real-time single-cell confocal microscopy using the methods described here, we have developed an effective protocol for investigating the mechanisms of IL-1β secretion and the testing of the hypothesis that IL-1β secretion requires membrane permeabilisation.

Keywords: Confocal microscopy; IL-1β Venus; IL-1β secretion; Lentiviral transduction; Real-time single-cell imaging.

MeSH terms

  • Animals
  • Cells, Cultured
  • Interleukin-1beta / metabolism*
  • Macrophages / cytology*
  • Macrophages / metabolism
  • Mice
  • Microscopy, Confocal
  • Signal Transduction
  • Single-Cell Analysis / methods*

Substances

  • IL1B protein, mouse
  • Interleukin-1beta