TY - JOUR
T1 - Spatial proteome profiling by immunohistochemistry-based laser capture microdissection and data-independent acquisition proteomics
AU - Huang, Peiwu
AU - Kong, Qian
AU - Gao, Weina
AU - Chu, Bizhu
AU - Li, Hua
AU - Mao, Yiheng
AU - Cai, Zongwei
AU - Xu, Ruilian
AU - Tian, Ruijun
N1 - Funding Information:
This study is supported by funding from National Natural Science Foundation of China (91953118), China State Key Basic Research Program Grants (2016YFA0501403 and 2016YFA0501404), Guangdong Provincial Fund for Distinguished Young Scholars (2019B151502050), Guangdong Provincial Natural Science Grant (2016A030312016), Shenzhen Innovation of Science and Technology Commission (JCYJ20170412154126026).
Publisher copyright:
© 2020 Elsevier B.V. All rights reserved.
PY - 2020/8/29
Y1 - 2020/8/29
N2 - Understanding the tumor heterogeneity through spatially resolved proteome profiling is important for biomedical research and clinical application. Laser capture microdissection (LCM) is a powerful technology for exploring local cell populations without losing spatial information. Conventionally, tissue sections are stained with hematoxylin and eosin (H&E) for cell-type identification before LCM. However, it generally requires experienced pathologists to distinguish different cell types, which limits the application of LCM to broad cancer research field. Here, we designed an immunohistochemistry (IHC)-based workflow for cell type-resolved proteome analysis of tissue samples. Firstly, targeted cell type was marked by IHC using antibody targeting cell-type specific marker to improve accuracy and efficiency of LCM. Secondly, to increase protein recovery from chemically crosslinked IHC tissues, we optimized a decrosslinking procedure to seamlessly combine with the integrated spintip-based sample preparation technology SISPROT. This newly developed approach, termed IHC-SISPROT, has comparable performance as H&E staining-based proteomic analysis. High sensitivity and reproducibility of IHC-SISPROT were achieved by combining with data independent acquisition proteomics. More than 3500 proteins were identified from only 0.2 mm2 and 12 μm thickness of hepatocellular carcinoma (HCC) tissue section. Furthermore, using 5 mm2 and 12 μm thickness of HCC tissue section, 6660 and 6052 protein groups were quantified from cancer cells and cancer-associated fibroblasts (CAFs) by the IHC-SISPROT workflow. Bioinformatic analysis revealed the enrichment of cell type-specific ligands and receptors and potentially new communications between cancer cells and CAFs by these signaling proteins. Therefore, IHC-SISPROT is a sensitive and accurate proteomic approach for spatial profiling of cell type-specific proteome from tissues.
AB - Understanding the tumor heterogeneity through spatially resolved proteome profiling is important for biomedical research and clinical application. Laser capture microdissection (LCM) is a powerful technology for exploring local cell populations without losing spatial information. Conventionally, tissue sections are stained with hematoxylin and eosin (H&E) for cell-type identification before LCM. However, it generally requires experienced pathologists to distinguish different cell types, which limits the application of LCM to broad cancer research field. Here, we designed an immunohistochemistry (IHC)-based workflow for cell type-resolved proteome analysis of tissue samples. Firstly, targeted cell type was marked by IHC using antibody targeting cell-type specific marker to improve accuracy and efficiency of LCM. Secondly, to increase protein recovery from chemically crosslinked IHC tissues, we optimized a decrosslinking procedure to seamlessly combine with the integrated spintip-based sample preparation technology SISPROT. This newly developed approach, termed IHC-SISPROT, has comparable performance as H&E staining-based proteomic analysis. High sensitivity and reproducibility of IHC-SISPROT were achieved by combining with data independent acquisition proteomics. More than 3500 proteins were identified from only 0.2 mm2 and 12 μm thickness of hepatocellular carcinoma (HCC) tissue section. Furthermore, using 5 mm2 and 12 μm thickness of HCC tissue section, 6660 and 6052 protein groups were quantified from cancer cells and cancer-associated fibroblasts (CAFs) by the IHC-SISPROT workflow. Bioinformatic analysis revealed the enrichment of cell type-specific ligands and receptors and potentially new communications between cancer cells and CAFs by these signaling proteins. Therefore, IHC-SISPROT is a sensitive and accurate proteomic approach for spatial profiling of cell type-specific proteome from tissues.
KW - Data-independent acquisition
KW - Immunohistochemistry
KW - Integrated sample preparation
KW - Laser capture microdissection
KW - Proteomics
UR - http://www.scopus.com/inward/record.url?scp=85087761130&partnerID=8YFLogxK
U2 - 10.1016/j.aca.2020.06.049
DO - 10.1016/j.aca.2020.06.049
M3 - Journal article
C2 - 32800117
AN - SCOPUS:85087761130
SN - 0003-2670
VL - 1127
SP - 140
EP - 148
JO - Analytica Chimica Acta
JF - Analytica Chimica Acta
ER -
