Multiplex immunofluorescence-guided laser capture microdissection for spatial transcriptomics of metastatic melanoma tissues

STAR Protoc. 2022 Dec 16;3(4):101698. doi: 10.1016/j.xpro.2022.101698. Epub 2022 Sep 22.

Abstract

We describe a pipeline for optimized and streamlined multiplexed immunofluorescence-guided laser capture microdissection allowing the harvest of individual cells based on their phenotype and tissue localization for transcriptomic analysis with next-generation RNA sequencing. Here, we analyze transcriptomes of CD3+ T cells, CD14+ monocytes/macrophages, and melanoma cells in non-dissociated metastatic melanoma tissue. While this protocol is described for melanoma tissues, we successfully applied it to human tonsil, skin, and breast cancer tissues as well as mouse lung tissues. For complete details on the use and execution of this protocol, please refer to Martinek et al. (2022).

Keywords: Cancer; Cell isolation; Immunology; Microscopy; RNAseq; Single cell.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Fluorescent Antibody Technique
  • Gene Expression Profiling / methods
  • Humans
  • Laser Capture Microdissection* / methods
  • Melanoma* / genetics
  • Melanoma* / surgery
  • Mice
  • Transcriptome / genetics