Agarose-Based Preparation of Human Precision-Cut Lung Slices

Begin with a resected human lung.

Insert a silicone tube into the main bronchus of the lung. Secure the tube, providing a pathway for introducing agarose into the tissue in the subsequent steps. 

Clamp other bronchi and vessels to prevent agarose leakage during filling.

Introduce pre-warmed low-gelling agarose mixed with media into the lung until it is fully inflated. 

Place the lung on ice for agarose polymerization, forming a solid gel matrix that provides stability to the tissue and maintains lung architecture during sectioning procedures.

After polymerization, cut the lung tissue into uniform slabs. Extract cylindrical tissue cores from the tissue slabs.

Using a tissue slicer filled with chilled buffer, slice the tissue core to obtain precision-cut lung slices.

Transfer the slices to a culture plate containing chilled media. Incubate to stabilize the tissue.

Finally, transfer the slices into a multi-well plate containing media. The prepared precision-cut lung slices are now ready for experiments.

To begin this procedure, cannulate the trachea by inserting the silicone tube, and fixing it with a clamp parallel to the tube so that the clamp squeezes the tissue together alongside