US 11,712,205 B2
High resolution intravital imaging and uses thereof
David Entenberg, Granite Springs, NY (US); John S. Condeelis, Bronx, NY (US); and Sonia E. Voiculescu, Bronx, NY (US)
Assigned to Albert Einstein College of Medicine, Bronx, NY (US)
Appl. No. 16/640,869
Filed by Albert Einstein College of Medicine, Bronx, NY (US)
PCT Filed Aug. 1, 2018, PCT No. PCT/US2018/044737
§ 371(c)(1), (2) Date Feb. 21, 2020,
PCT Pub. No. WO2019/040244, PCT Pub. Date Feb. 28, 2019.
Claims priority of provisional application 62/548,455, filed on Aug. 22, 2017.
Prior Publication US 2020/0352516 A1, Nov. 12, 2020
Int. Cl. A61B 5/00 (2006.01); A61B 5/08 (2006.01); G02B 21/36 (2006.01)
CPC A61B 5/704 (2013.01) [A61B 5/0068 (2013.01); A61B 5/0071 (2013.01); A61B 5/08 (2013.01); A61B 5/418 (2013.01); A61B 5/4222 (2013.01); A61B 5/4872 (2013.01); G02B 21/367 (2013.01); A61B 2503/42 (2013.01)] 29 Claims
OG exemplary drawing
 
1. A method of high-resolution intravital imaging of lung tissue in a subject using a microscope, comprising:
a) exposing the lung tissue in the subject by anesthetizing the subject and making a circular incision through the skin and adjacent ribs of the subject, lateral to the sternum and superior to the last floating rib;
b) attaching a window frame to the subject so that the lung tissue is viewable through the window; wherein the window frame is attached by securing the window frame in the circular incision so that the adjacent ribs are positioned in a groove of the window frame; wherein the window frame comprises fiducial marks; and wherein the window frame is adhered to the lung tissue by raising the window frame away from the lung tissue, dispensing a quick-setting adhesive on the underside of the window frame, and expanding the lung to make contact with the window frame until the adhesive sets, suturing tissue around the window frame to provide a complete surgical closure, and sealing the thoracic cavity by applying an adhesive to a cover glass and adhering it to the expanded lung tissue and the inner recess of the window frame;
c) following recovery of the subject from surgery, attaching the window frame to a microscope stage with a window fixturing plate that is movable relative to the optical axis of the microscope in at least x- and y-directions, wherein the window frame is attached to the microscope stage and stabilized for imaging by placing a fixturing plate, comprising portions defining a slotted recess sized to accommodate the window frame, between skin of the subject and an outside lip of the window frame, and placing the subject on the stage of the microscope; and
d) acquiring through the window high-magnification, high-resolution overlapping images of the lung tissue in a mosaic pattern and stitching the images together to produce a low-magnification, high-resolution overview image of the lung tissue that preserves subcellular resolution, thereby obtaining high-resolution intravital images of the lung tissue in a subject.