This article describes a technique for high-resolution intravital microscopy using an implanted pancreatic imaging window. This method allows for long-term visualization of pancreatic islets in live animals, enhancing the study of pancreatic functions.
High-resolution intravital microscopy, via an implanted pancreatic imaging window, facilitates long-term, wide-area cellular-level visualization of the pancreatic islets in a live animal.
For pancreatic imaging window implantation, first, position an anesthetized mouse in a left lateral position. Next, fix a probe to monitor the mouse’s temperature and regulate its temperature using a heating pad. Now, shave the left flank and sterilize the area.
Make a surgical incision and dissect the abdominal muscles. Insert appropriate sutures in the incision margin to affix the imaging window frame in position following implantation. Further, expose and exteriorize the spleen, and identify the connected pancreas.
Next, overlay the pancreas connected with the spleen on the plate of the customized pancreatic imaging window, with the spleen positioned in the open space of the window. This positioning of the pancreas effectively isolates it from the neighboring bowel movements, reducing the motion-induced artifacts during imaging. Carefully apply cyanoacrylate glue to the margin of the imaging window, avoiding the organs.
Mount a cover glass over the margin of the imaging window. Tighten the sutures around the imaging window to secure the window in place. The mouse with the implanted pancreatic imaging window is ready for intravital pancreatic imaging.
Begin by preparing the surgical platform, and disinfecting the surfaces with 70% ethanol. After anesthetization, use a rectal probe with a homeothermic controlled heating pad to monitor the body temperature.
Remove hair from the left flank of the mouse and apply alcohol and iodine alternatively, rotating from the incision site to the outer surface, never going back to the incision site and ending with iodine. Then make a 1.5-centimeter incision on the left flank of the mouse dissecting the skin and the muscle.
Use a black or nylon 4-0 suture to perform a purse-string suture in the incision margin. Pool the spleen carefully with ring forceps, and identify the pancreas. Placing the window at the flank of the mouse, pass the spleen and the pancreas through the open space of the window.
Then, gently place the pancreas on the plate of the imaging window while placing the spleen on the open space of the window. Use a 31 gauge catheter needle to apply drops of N-butyl cyanoacrylate glue to the margin of the imaging window, ensuring a minimal amount of glue is applied.
Gently apply a 12-millimeter round cover glass to the margin of the imaging window. Then, hold the suture loop to fit into the lateral groove of the window and tie it three times. Finally, to prevent the interruption of these tight stitches when the mice are awake, cut the maximal proximal site of the tie.
繁體中文
