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Cell Biology JoVE (Open Access) Citable · DOI

High Resolution 3D Imaging of the Human Pancreas Neuro-insular Network

DOI: 10.3791/56859-v
What you'll learn
  • ✓ Perform passive optical clearing on human pancreas tissue sections
  • ✓ Execute multi-round immunofluorescence staining for neural markers
  • ✓ Acquire and interpret 3D images of neuro-insular networks
Protocol

Biopharma Insights Here, we present a protocol to image human pancreas sections in three dimensions (3D) using optimized passive clearing methods. This manuscript demonstrates these procedures for passive optical clearing followed by multiple immunofluorescence staining to identify key elements of the autonomic and sensory neural networks innervating human islets.

Difficulty
advanced
Total time
~5–7 days (including fixation, clearing, staining, and imaging)
Biosafety
BSL-1

Steps

1
Fix pancreas tissue and embed in hydrogel

Prepare human pancreas sections through fixation and hydrogel embedding to preserve tissue architecture and enable subsequent clearing and staining procedures.

▶ 00:35
2
Clear tissue using passive optical clearing methods

Apply optimized passive clearing protocols to increase tissue transparency and improve light penetration for 3D imaging of deep neural structures.

▶ 03:37
3
Perform iterative immunofluorescence staining rounds

Execute multiple rounds of antibody staining to label autonomic, sensory, and supporting neural cell markers within the pancreatic islet network.

▶ 04:38
4
Mount cleared samples for 3D imaging acquisition

Prepare stained tissue for high-resolution confocal or light-sheet microscopy to capture volumetric data of the neuro-insular network.

▶ 07:03
5
Visualize and reconstruct 3D neural networks

Analyze acquired image stacks to resolve Schwann cells, autonomic neurons, and sensory innervation patterns within human pancreatic islets in three dimensions.

▶ 08:30
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