Tissue Immunostaining for Imaging Mass Cytometry

Abstract

Source:
Jake Griner¹, Arwen Wedgeworth², Brenton Puckett², Kacey Idouchi¹, Slivia Guglietta², Carsten Krieg<sup>1

1</sup>Department of Pathology and Laboratory Medicine, Medical University of South Carolina.

²Department of Regenerative Medicine and Cell Biology, Medical University of South Carolina.

The video describes the preparation of a tissue sample for imaging mass cytometry. The tissue is first processed to unmask the antigen epitopes, permeabilize the cells, and block non-specific antibody binding sites. Then, the tissue is incubated with a metal-conjugated antibody cocktail and a nuclear stain. Finally, the treated tissue sample is imaged using an optical scanner.

Protocol

1. Tissue pretreatment:

For Fresh Frozen tissues:

1. Place a glass Coplin jar in the -20°C freezer for at least one hour before use.
2. Remove the slides from the -80°C freezer and place them into the glass Coplin jar in the -20°C freezer for at least 15 minutes.
3. Under the fume hood, use an ampule cutter to open the 16% paraformaldehyde (PFA) glass ampule. Discard the broken glass appropriately.
4. Add 10 mL of 16% PFA and 30 mL phosphate-buffered saline (PBS) to a plastic Coplin jar. Ensure that the PBS is pre-chilled to 4°C.
5. Transfer this solu....