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Atomic force microscopy of peritoneal macrophages after particle phagocytosis
1University of Tübingen, Institute of Physiology, Germany.
Abstract:
The atomic force microscope was used to image peritoneal macrophages after phagocytosis of latex beads with 0.45 microns in diameter and of zymosan particles. The rigidity of the phagocytosed material allowed to image the live membrane at forces below 2 nN. Repeated scanning of the membrane unavoidably caused the protrusion of the beads and increased their virtual height. The influence of fixation by glutaraldehyde on the image and the corresponding force vs. distance curves were analyzed and compared. Short treatment with Triton X-100 enabled us to identify intracellular components, such as embedded latex beads, cell nucleus and cytoskeletal strands. The data demonstrate that it is possible to image living cells if they are bolstered by stiff material.
Insights
Atomic force microscopy can image live peritoneal macrophages by using stiff phagocytosed materials like latex beads. This method allows visualization of cell membranes and intracellular components without damaging the cells.
Area of Science:
- Cell biology
- Biophysics
- Microscopy
Background:
- Peritoneal macrophages are crucial immune cells.
- Imaging live cells presents challenges due to membrane fragility.
- Phagocytosis is a key cellular process involving particle uptake.
Purpose of the Study:
- To investigate the feasibility of imaging live peritoneal macrophages using atomic force microscopy (AFM).
- To assess the role of phagocytosed stiff materials in stabilizing live cell membranes for AFM imaging.
- To analyze the effects of fixation and detergent treatment on AFM imaging of macrophages.
Main Methods:
- Atomic force microscopy (AFM) was employed to image live peritoneal macrophages.
- Macrophages were induced to phagocytose latex beads (0.45 microns) and zymosan particles.
- Imaging was performed at low forces (< 2 nN) to minimize membrane damage.
- Effects of glutaraldehyde fixation and Triton X-100 treatment were analyzed.
Main Results:
- Stiff phagocytosed materials (latex beads, zymosan) enabled stable imaging of live macrophage membranes.
- Repeated AFM scanning led to bead protrusion and increased virtual height.
- Glutaraldehyde fixation altered image characteristics and force curves.
- Triton X-100 treatment allowed visualization of intracellular structures like the nucleus and cytoskeleton.
Conclusions:
- Atomic force microscopy can successfully image live peritoneal macrophages when supported by rigid phagocytosed particles.
- The rigidity of ingested material is key to maintaining membrane integrity during AFM analysis.
- This technique offers a method for studying live cell dynamics and internal structures non-invasively.