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Development of a sample preparation protocol for 3D kidney ultrastructural analysis and immunolabeling by light microscopy

Award Number: R43DK137685
ORGANIZATION: NATIONAL INSTITUTE OF DIABETES & DIGESTIVE & KIDNEY DISEASES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 09/15/2023
PERIOD OF PERFORMANCE END DATE: 08/31/2024

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Development of a sample preparation protocol for 3D kidney ultrastructural analysis and immunolabeling by light microscopy - Abstract The success of cohort studies such as the NIDDK Kidney Precision Medicine Project (KPMP) with its massive amount of samples depends on the cost and time-efficient evaluation of kidney biopsies at multiple scales. Assessing the structure of kidney tissue at the nanoscale (~10 nm) and evaluating the 3-D distribution of protein biomarkers in that context is, however, currently severely limited as it depends on expensive, time-consuming and extremely complex correlative light and electron microscopy (CLEM) as the only suitable tool. Panluminate has developed a proprietary new tissue expansion technology we call pan-Expansion Microscopy (pan-ExM), that directly addresses this challenge. By combining ~25-fold linear expansion of biological samples with novel bulk (pan) staining of amino acids, pan-ExM is capable of mapping the 3-D nanoscale distribution of specific biomarkers in the context of kidney ultrastructure. Hallmark ultrastructural features such as podocyte foot processes and cristae in mitochondria can now be resolved by their characteristic pan-staining pattern, analogous to heavy-metal stains in electron microscopy (EM). In contrast to EM/CLEM, however, pan-ExM is an all-optical method that allows researchers to interrogate tissue samples using standard confocal microscopes; this offers important advantages in terms of time, cost, complexity and sample preservation versus EM alternatives. Our proposed Phase I project is to develop and validate pan-ExM for kidney samples. Specifically we will: (1) optimize and validate a new antibody-labeling pan-ExM protocol in freshly fixed kidney tissue; and (2) develop and validate a novel ~25x tissue expansion protocol for archived FFPE kidney biopsies. This work is designed to lay the foundation for a Phase II project that will establish pan-ExM as a robust and powerful imaging technology in the kidney field by enabling every researcher in nephrology to locate multiple specific protein biomarkers within their 3D contextual compartments at the ~10 nm scale. Our long-term ambition is for pan-ExM to become a core imaging technology for kidney tissue atlases as well as a preferred technique for targeted research and diagnostic applications, thereby paving the road to precision medicine for nephrology.


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: 2025 ( Subtotal = -$28 )
2025	2023	PANLUMINATE INC	101 COLLEGE STREET, SUITE 210	NEW HAVEN	CT	06510	SOUTH CENTRAL CT	USA	Diabetes, Digestive, and Kidney Diseases Extramural Research	000	1	1/21/2025	NEW	-$28
														Subtotal = -$28

Issue Date FY: 2023 ( Subtotal = $274,419 )
2023	2023	PANLUMINATE INC	123 YORK ST	NEW HAVEN	CT	06511	SOUTH CENTRAL CT	USA	Diabetes, Digestive, and Kidney Diseases Extramural Research	000	1	9/15/2023	NEW	$274,419
														Subtotal = $274,419

Grand Total All Awards = $274,391

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Development of a sample preparation protocol for 3D kidney ultrastructural analysis and immunolabeling by light microscopy

Award Number: R43DK137685

ORGANIZATION: NATIONAL INSTITUTE OF DIABETES & DIGESTIVE & KIDNEY DISEASES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

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

PERIOD OF PERFORMANCE START DATE: 09/15/2023

PERIOD OF PERFORMANCE END DATE: 08/31/2024

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