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Single Cell Proteoform Profiling using Top-Down Mass Spectrometry

Award Number: R00AI183290
ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 02/06/2026
PERIOD OF PERFORMANCE END DATE: 01/31/2028

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Single Cell Proteoform Profiling using Top-Down Mass Spectrometry - Project Summary/Abstract Proteoforms are molecular forms of proteins in cells and tissues containing site-specific sequence variations and post-translational modifications. Proteoforms effectively describe cell phenotypes and provide important implications in disease mechanisms. Recent studies in immunobiology and disease pathology have emphasized intact proteoform characterization without enzymatic digestion in bottom-up approaches. Current mass-spectrometry-based top-down proteomics technologies fall short in capturing the complex proteome in small biological samples such as single cells. This proposal generates a suite of novel technologies for high-throughput omics- scale single cell proteoform profiling using innovative instrumentation, bioinformatics and high- throughput strategies. I will use human kidney cells as a model biological system in this proposal, and the approaches are generalizable to different cell types. I have recently developed a technology employing localized proteoform sampling coupled to single-molecule mass spectrometry to directly image and identify intact proteoforms in tissue sections (Su et al., 2022, Sci. Adv.). In Aim 1, I will expand the proteome detection and identification capabilities in this technology using innovative instrumentation, sampling and data acquisition algorithms. Together these will curate a knowledgebase serving as a proteoform library for kidney single cell proteoform profiling. In Aim 2, I will develop a novel platform leveraged for profiling of single cells dissociated from human kidney biopsies. I will develop a single cell preparation protocol for maximizing proteoform detection in kidney parenchymal and immune cells. I will also develop a bioinformatics approach tailored for proteoform identification in single cell datatype and discover proteoform signatures that differentiate cell types. In Aim 3, I will address the limitation in rare cell profiling by developing a series of high-throughput strategies including high-speed sampling and microarray cell patterning. These new technologies will unravel proteoform landscapes and signatures in rare kidney-resident innate immune cells (e.g., macrophages and dendritic cells) for the discovery of new cell populations that can be used as therapeutic targets and diagnostic tools for inflammatory diseases. My mentoring team consists of Dr. Neil Kelleher (mentor), a world-renowned protein biochemist, and Dr. Satish Nadig (co-mentor), a leading expert in kidney immunobiology. The proposed research is a substantial technological advancement in single-cell proteomics and sets a solid foundation for the pursuit of my independent career. The proposed research also well aligns with my long term research interest in developing enabling analytical technologies for biomedical science community with a special interest in the human innate immune system.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2026 ( Subtotal = $249,000 )
2026	2026	REGENTS OF THE UNIVERSITY OF CALIFORNIA AT RIVERSIDE	200 UNIVERSTY OFC BUILDING	RIVERSIDE	CA	92521	RIVERSIDE	USA	Allergy and Infectious Diseases Research	000	3	2/6/2026	EXTENSION WITH OR WITHOUT FUNDS	$249,000
														Subtotal = $249,000

Grand Total All Awards = $249,000

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Single Cell Proteoform Profiling using Top-Down Mass Spectrometry

Award Number: R00AI183290

ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

PERIOD OF PERFORMANCE START DATE: 02/06/2026

PERIOD OF PERFORMANCE END DATE: 01/31/2028

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