Filariasis Research Reagent Resource Center

FILARIASIS RESEARCH REAGENT RESOURCE CENTER:

University of Wisconsin Oshkosh

STANDARD OPERATING PROCEDURE

TITLE: Surgical Transplantation of Adult Acanthocheilonemaviteae into Gerbils

SOP NUMBER: A. vit. 7

1.PURPOSE

The purpose of this document is to describe the procedure used when subcutaneously transplanting adult Acanthocheilonemaviteae into recipient gerbils.

2.MATERIALS

2.1Anesthesia of choice (i.e., isoflurane)

2.2Source of heat to provide thermal support for the gerbil during surgery

2.3Approved pain medication for gerbil (i.e.,Ketofen®; diluted to 3 mg/mL; 0.1 mL given SQ)

2.4Non-medicated eye lubricant for gerbil

2.5Sterile plastic transfer pipettes

2.6 Twosterile handmade dissection tools (27 gauge needles bent at a 45º angle and placed on the end of a cotton tip applicator). These ‘worm hooks’ can be used to move adult worms.

2.7Sterile surgical pack to include scalpel, scissors, needle holders, hemostats as needed by surgeon.

2.870% ethanol

2.9Sterile cotton tipped applicators

2.10Sterile drapes or towels

2.11Suture with attached needle

2.12Surgical light

2.13Sterile gloves for surgeon

2.14Small hair clippers

2.15Surgical skin glue

2.16Adult A. viteaecultured in a plastic Petri dish until ready for the transplant procedure

3.0PROCEDURE

3.1Anesthetize the gerbil with isoflurane. Use 5% isoflurane to induce the animal and 2.5-3.5% for maintenance anesthesia during the procedure. Isoflurane is used as a mixture with oxygen (100%) at a flow rate of 1 L/min. Any suitable alternative that produces a surgical plane of anesthesia can be substituted for isoflurane but alternatives must first be approved for use on the IACUC research protocol.

3.2Position the animal in ventral recumbency on the source of thermal support.

Note:Prior to ever using the heat source you choose, be sure to test it to ensure it does not get too hot, which could result in death of the animal or serious burns. A stockinet sleeve filled with uncooked rice warmed briefly in the microwave (i.e., 30-45 seconds) works well, as it holds heat well and allows for proper positioning of the animal. The rice filled stockinet can also be autoclaved between uses.

3.3Prepare the gerbilby clipping the hair off of the animal’s back, starting near the back of the skull and stopping about 1 inch behind the should blades. Swab the skin at that location with two rounds of 70% ethanol and a sterile cotton tipped applicator.

3.4Apply eye lubricant to both eyes of the anesthetized animal.

3.5Drape the incision site with a sterile fenestrated drape or towel.

3.6Make a small incision through the skin layer between the shoulder blades, taking care not to cut any muscle or other underlying structures. Use a blunt probe or blunt scissors to bluntly dissect the underlying connective tissue to create a small ‘pocket’. This pocket will receive the transplanted worms.

3.7Manipulate the desired number of worms in the Petri dish into a clump. Insert the worms into the incision. To do so, have the surgeon hold the incision open with their surgical instruments and an assistant place the worms in the subcutaneous pocket using sterile worm hooks or sterile transfer pipette. The hooks often work well for a clump of worms, while the pipette is better suited for one or two worms. Be sure not to poke the worms with the hooks, but rather use them to scoop the clump of worms.

3.8The surgeon should ensure that the worms are at least a few mm away from the incision so they are not caught in the suture while the incision is being closed.

3.9Close the incision with three or more sutures, depending on the length of the incision. Use a few small drops of skin glue along the suture line to help keep it closed. It is not uncommon for the gerbils to immediately attempt to remove their sutures upon recovery from anesthesia.

3.10Administer pain medication. This is often required by IACUC and will help reduce the likelihood of the animals removing their sutures.

3.11Monitor the gerbil(s) closely during recovery from anesthesia.

3.12Time to Completion: 45-60 minutes per gerbil

Last revision: 3/5/14 SS & LMA. vit. 7