A Dissection Method to Harvest Murine Popliteal Draining Lymph Nodes for Ex Vivo Imaging

Abstract

Source: Rezende, R. M., et al. Visualizing Lymph Node Structure and Cellular Localization using Ex-Vivo Confocal Microscopy. J. Vis. Exp. (2019)

This video demonstrates a dissection technique to harvest popliteal draining lymph nodes from the mouse hind legs. Lymph nodes play a critical role in bridging innate and adaptive immunity, and harvesting intact lymph nodes — with a preserved tissue structure and cellular viability — can help study local and systemic immune responses.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board. 1. Surgery procedure to remove the popliteal draining lymph node Euthanize mice using CO2 asphyxiation followed by cervical dislocation. Immobilize mice at a prone position on the acrylic stage with adhesive tape and apply mineral oil with a cotton swab in the calf and knee (Figure 1A-D</strong…

Representative Results

Figure 1: Popliteal lymph node preparation. (A, B) Subcutaneous injection of FACS antibody master mix into the paw pad. (C, D) 3 h after the injection, euthanize the mouse, immobilize the mouse on an acrylic plate with adhesive tape (C), and apply mineral oil to the abdominal skin to prevent fur deposition around the incision (D). (E) Perform a …

Materials

Cellview culture dish	Greiner-Bio	627861	35×10 mm with glass bottom
Kimwipes	Kimtech Science Brand	7557	size 21 x 20 cm / 100 sheets per box
Microsurgery curved forceps	WEP Surgical Instruments	Custom-made	12.5 cm
Microsurgery curved scissors	WEP Surgical Instruments	Custom-made	11.5 cm