Highly efficient and robust π-FISH rainbow for multiplexed in situ detection of diverse biomolecules

Yingfeng Tao; Xiaoliu Zhou; Leqiang Sun; Da Lin; Huaiyuan Cai; Xi Chen; Wei Zhou; Bing Yang; Zhe Hu; Jing Yu; Jing Zhang; Xiaoqing Yang; Fang Yang; Bang Shen; Wenbao Qi; Zhenfang Fu; Jinxia Dai; Gang Cao

doi:10.1038/s41467-023-36137-4

Highly efficient and robust π-FISH rainbow for multiplexed in situ detection of diverse biomolecules

Nat Commun. 2023 Jan 27;14(1):443. doi: 10.1038/s41467-023-36137-4.

Authors

Yingfeng Tao^#^{1

2}, Xiaoliu Zhou^#^{1

2}, Leqiang Sun^{1

2}, Da Lin^{1

2}, Huaiyuan Cai^{1

2}, Xi Chen^{1

2}, Wei Zhou^{1

2}, Bing Yang^{1

2}, Zhe Hu¹, Jing Yu³, Jing Zhang⁴, Xiaoqing Yang⁵, Fang Yang⁶, Bang Shen^{1

2

7}, Wenbao Qi^{8

9}, Zhenfang Fu¹⁰, Jinxia Dai^{11

12}, Gang Cao^{13

14

15}

Affiliations

¹ State Key Laboratory of Agricultural Microbiology, Hubei Hongshan Laboratory, Huazhong Agricultural University, 430070, Wuhan, China.
² College of Veterinary Medicine, Huazhong Agricultural University, 430070, Wuhan, China.
³ Department of Blood Transfusion, Wuhan hospital of Traditional Chinese and Western Medicine, Huazhong University of Science and Technology, 430070, Wuhan, China.
⁴ Department of the 1st Thoracic Medical Oncology, Hubei Cancer Hospital, Tongji Medical College, Huazhong University of Science and Technology, 430070, Wuhan, China.
⁵ Hospital of Huazhong Agricultural University, 430070, Wuhan, China.
⁶ National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, 430070, Wuhan, China.
⁷ Key Laboratory of Preventive Medicine in Hubei Province, 430070, Wuhan, Hubei Province, China.
⁸ College of Veterinary Medicine, South China Agricultural University, 510642, Guangzhou, China.
⁹ African Swine Fever Regional Laboratory of China, Guangzhou, China.
¹⁰ Departments of Pathology, College of Veterinary Medicine, University of Georgia, Athens, GA, 30602, USA.
¹¹ State Key Laboratory of Agricultural Microbiology, Hubei Hongshan Laboratory, Huazhong Agricultural University, 430070, Wuhan, China. jxdai@mail.hzau.edu.cn.
¹² College of Veterinary Medicine, Huazhong Agricultural University, 430070, Wuhan, China. jxdai@mail.hzau.edu.cn.
¹³ State Key Laboratory of Agricultural Microbiology, Hubei Hongshan Laboratory, Huazhong Agricultural University, 430070, Wuhan, China. gcao@mail.hzau.edu.cn.
¹⁴ College of Veterinary Medicine, Huazhong Agricultural University, 430070, Wuhan, China. gcao@mail.hzau.edu.cn.
¹⁵ College of Biomedicine and Health, Huazhong Agricultural University, 430070, Wuhan, China. gcao@mail.hzau.edu.cn.

^# Contributed equally.

Abstract

In the unprecedented single-cell sequencing and spatial multiomics era of biology, fluorescence in situ hybridization (FISH) technologies with higher sensitivity and robustness, especially for detecting short RNAs and other biomolecules, are greatly desired. Here, we develop the robust multiplex π-FISH rainbow method to detect diverse biomolecules (DNA, RNA, proteins, and neurotransmitters) individually or simultaneously with high efficiency. This versatile method is successfully applied to detect gene expression in different species, from microorganisms to plants and animals. Furthermore, we delineate the landscape of diverse neuron subclusters by decoding the spatial distribution of 21 marker genes via only two rounds of hybridization. Significantly, we combine π-FISH rainbow with hybridization chain reaction to develop π-FISH+ technology for short nucleic acid fragments, such as microRNA and prostate cancer anti-androgen therapy-resistant marker ARV7 splicing variant in circulating tumour cells from patients. Our study provides a robust biomolecule in situ detection technology for spatial multiomics investigation and clinical diagnosis.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Humans
In Situ Hybridization, Fluorescence / methods
Male
MicroRNAs* / genetics
Nucleic Acids*
Prostatic Neoplasms* / diagnosis
Prostatic Neoplasms* / genetics

Substances

MicroRNAs
Nucleic Acids