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Cyclic Microchip Assay for Measurement of Hundreds of Functional Proteins in Single Neurons

Liwei Yang, Avery Ball, Jesse Liu, Tanya Jain, Yue-Ming Li, Jun Wang
doi: https://doi.org/10.1101/2021.06.06.447288
Liwei Yang
1Multiplex Biotechnology Laboratory, Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY 11794
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Avery Ball
1Multiplex Biotechnology Laboratory, Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY 11794
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Jesse Liu
1Multiplex Biotechnology Laboratory, Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY 11794
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Tanya Jain
2Chemical Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
3Programs of Neurosciences, Weill Graduate School of Medical Sciences of Cornell University, New York, NY, USA
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Yue-Ming Li
2Chemical Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY, USA
3Programs of Neurosciences, Weill Graduate School of Medical Sciences of Cornell University, New York, NY, USA
4Programs of Pharmacology, Weill Graduate School of Medical Sciences of Cornell University, New York, NY, USA
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Jun Wang
1Multiplex Biotechnology Laboratory, Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY 11794
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  • For correspondence: Jun.wang.5@stonybrook.edu
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Abstract

Proteins are responsible for nearly all cell functions throughout cellular life. To date, the molecular functions of hundreds of proteins have been studied as they are critical to cellular processes. Those proteins are varied dramatically at different statuses and differential stages of the cells even in the same tissue. The existing single-cell tools can only analyze dozens of proteins and thus have not been able to fully characterize a cell yet. Herein, we present a single-cell cyclic multiplex in situ tagging (CycMIST) technology that affords the comprehensive functional proteome profiling of single cells. It permits multiple, separate rounds of multiplex assays of the same single cells on a microchip where each round detects 40-50 proteins. A decoding process is followed to assign protein identities and quantify protein detection signals. We demonstrate the technology on a neuron cell line by detecting 182 proteins that includes surface makers, neuron function proteins, neurodegeneration markers, signaling pathway proteins and transcription factors. Further study on 5XFAD mouse, an Alzheimer’s Disease (AD) model, cells validate the utility of our technology which reveals the deep heterogeneity of brain cells. Through comparison with control mouse cells, the differentially expressed proteins in the AD mouse model have been detected. The single-cell CycMIST technology can potentially analyze the entire functional proteome spectrum, and thus it may offer new insights into cell machinery and advance many fields including systems biology, drug discovery, molecular diagnostics, and clinical studies.

Competing Interest Statement

YML is a co-inventor of intellectual property (assay for gamma secretase activity and screening method for gamma secretase inhibitors) owned by MSKCC and licensed to Jiangsu Continental Medical Development.

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC-ND 4.0 International license.
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Posted June 07, 2021.
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Cyclic Microchip Assay for Measurement of Hundreds of Functional Proteins in Single Neurons
Liwei Yang, Avery Ball, Jesse Liu, Tanya Jain, Yue-Ming Li, Jun Wang
bioRxiv 2021.06.06.447288; doi: https://doi.org/10.1101/2021.06.06.447288
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Cyclic Microchip Assay for Measurement of Hundreds of Functional Proteins in Single Neurons
Liwei Yang, Avery Ball, Jesse Liu, Tanya Jain, Yue-Ming Li, Jun Wang
bioRxiv 2021.06.06.447288; doi: https://doi.org/10.1101/2021.06.06.447288

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