NIAID Schistosomiasis Resource Center

Perfusion of adult worms from mice

Author Fred Lewis

Introduction

One of the more common procedures in a schistosome life cycle laboratory is the collection of adult worms from the definitive mammalian host. In a bisexual worm infection in the mouse, adult S. mansoni typically reside in the mesenteric veins, from which they can be harvested by perfusion of the portal venous system.

Equipment

Peristaltic perfusion pump (e.g., Masterflex Console Drive/Cole-Palmer Inst. Co.)

Silicone tubing (e.g., Masterflex 96420-14) fitted with a 20-gauge needle

Foot-pedal for peristaltic pump (e.g., Treadlite II/Linemaster Switch Corp.)

Materials and Reagents

S. mansoni-infected mice

Sodium pentobarbital solution (65 mg/ml) containing heparin (10,000 U/ml)

Perfusion fluid (0.85% sodium chloride + 0.75% sodium citrate)

Small sharp scissors for dissecting mice

Procedure

· Euthanize the mouse with an intraperitoneal injection of 0.3 ml pentobarbital/heparin solution.

· Dissect the mouse so that its abdominal and thoracic cavities are opened.

· Carefully sever the ribs near the spinal column of the (mouse’s) left half of the thoracic cavity.

· Make a small slit in the hepatic portal vein, then insert the 20 gauge needle into the descending aorta.

· Pump perfusion fluid through the needle, and collect the perfusate in a container.

We use 7” x 11”x 3” plastic pans.

Follow-up comments/recommendations

We highly recommend using a foot-pedal to operate the peristaltic pump. This greatly facilitates the perfusion process by leaving both hands free to perform the procedure. Perfusing out every adult worm without taking additional steps is difficult, since some worms may still be trapped in the mesenteries, and others may have been swept into the liver. One may see worms trapped in the liver by removing the liver after perfusion and compressing it between two glass plates. If every worm must be accounted for, one must examine the mesenteries using a dissecting microscope.

References

Duvall, R.H. and DeWitt, W.B. 1967. An improved perfusion technique for recovering adult schistosomes from laboratory animals. The American Journal of Tropical Medicine and Hygiene 16: 483-486.

Lewis, F. A. 1999. Schistosomiasis, in Current Protocols in Immunology, John Wiley and Sons, Inc., (R. Coico, Ed) Vol III, 19.1.1 – 19.1.28.

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		Perfusion of adult worms from mice Author Fred Lewis Introduction One of the more common procedures in a schistosome life cycle laboratory is the collection of adult worms from the definitive mammalian host. In a bisexual worm infection in the mouse, adult S. mansoni typically reside in the mesenteric veins, from which they can be harvested by perfusion of the portal venous system. Equipment Peristaltic perfusion pump (e.g., Masterflex Console Drive/Cole-Palmer Inst. Co.) Silicone tubing (e.g., Masterflex 96420-14) fitted with a 20-gauge needle Foot-pedal for peristaltic pump (e.g., Treadlite II/Linemaster Switch Corp.) Materials and Reagents S. mansoni-infected mice Sodium pentobarbital solution (65 mg/ml) containing heparin (10,000 U/ml) Perfusion fluid (0.85% sodium chloride + 0.75% sodium citrate) Small sharp scissors for dissecting mice Procedure · Euthanize the mouse with an intraperitoneal injection of 0.3 ml pentobarbital/heparin solution. · Dissect the mouse so that its abdominal and thoracic cavities are opened. · Carefully sever the ribs near the spinal column of the (mouse’s) left half of the thoracic cavity. · Make a small slit in the hepatic portal vein, then insert the 20 gauge needle into the descending aorta. · Pump perfusion fluid through the needle, and collect the perfusate in a container. We use 7” x 11”x 3” plastic pans. Follow-up comments/recommendations We highly recommend using a foot-pedal to operate the peristaltic pump. This greatly facilitates the perfusion process by leaving both hands free to perform the procedure. Perfusing out every adult worm without taking additional steps is difficult, since some worms may still be trapped in the mesenteries, and others may have been swept into the liver. One may see worms trapped in the liver by removing the liver after perfusion and compressing it between two glass plates. If every worm must be accounted for, one must examine the mesenteries using a dissecting microscope. References Duvall, R.H. and DeWitt, W.B. 1967. An improved perfusion technique for recovering adult schistosomes from laboratory animals. The American Journal of Tropical Medicine and Hygiene 16: 483-486. Lewis, F. A. 1999. Schistosomiasis, in Current Protocols in Immunology, John Wiley and Sons, Inc., (R. Coico, Ed) Vol III, 19.1.1 – 19.1.28.