The Day‐3 Thymectomy Model for Induction of Multiple Organ‐Specific Autoimmune Diseases

Elisabeth Suri‐Payer1, Kevin Wei1, Kenneth Tung2

1 National Institute of Allergy and Infectious Diseases National Institute of Health, Bethesda, Maryland, 2 University of Virginia, Charlottesville, Virginia
Publication Name:  Current Protocols in Immunology
Unit Number:  Unit 15.16
DOI:  10.1002/0471142735.im1516s32
Online Posting Date:  May, 2001
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Abstract

Autoimmune diseases of the ovary and the stomach are major components of the murine autoimmune polyendocrinopathy syndrome that occurs following experimental perturbation of the normal immune system. Several methods of perturbation can lead to autoimmune disease, and the disease occurs in multiple organs in association with autoantibodies to tissue‐specific antigen of the respective organs. The autoimmune diseases develop in a variety of laboratory mouse strains that are not prone to spontaneous autoimmune diseases. This unit describes the induction of autoimmune disease of the ovary and the stomach in mice by thymectomy at the age of day 3 (day‐3 thymectomy) and methods for evaluating disease histologically and serologically.

     
 
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Table of Contents

  • Basic Protocol 1: Induction of Autoimmune Disease of the Ovary and Stomach by Day‐3 Thymectomy
  • Support Protocol 1: Histopathologic Evaluation of Autoimmune Disease of the Ovary and Autoimmune Gastritis
  • Support Protocol 2: Detection of Serum Autoantibody to Ovarian and Gastric Antigens Using Indirect Immunofluorescence
  • Reagents and Solutions
  • Commentary
  • Figures
  • Tables
     
 
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Materials

Basic Protocol 1: Induction of Autoimmune Disease of the Ovary and Stomach by Day‐3 Thymectomy

  Materials
  • Metofane (Mallinckrodt Veterinary)
  • Breeding pair of disease‐susceptible mouse strain (Table 15.16.1) with 3‐day‐old pups
  • Wescodyne or Betadine (Purdue‐Frederick)
  • 70% alcohol
  • Heat lamp
  • Superglue (Sparco 01264, UHU All Purpose Power Super Glu; Eberhard Farber)
  • 4 × 4–in sterile gauze
  • Glass jar with metal top
  • 100‐mm petri dishes and small plastic containers with mouse bedding to hold the pups postoperatively
  • Styrofoam or wax board with 1 round and 3 flat rubber bands to restrain pups
  • Fiber‐optic cool light source (Dolan Jenner)
  • Sterile cotton applicators (Pur‐wraps; Hardwood Product)
  • Binocular dissecting microscope
  • Surgical instruments (autoclaved) including:
  •  Fine McPherson‐Vannas (microdissecting) scissors (Roboz; 2 pairs)
  •  Fine Foerster (microdissecting) forceps (Roboz)
  • 5¾ in. (∼14.25 cm)–long glass Pasteur pipets with 2‐mm‐wide tips, autoclaved
  • Electric vacuum pump with adjustable suction strengths connected to pipets via long vacuum tubing, interrupted by a “waste” container to collect the thymic lobes

Support Protocol 1: Histopathologic Evaluation of Autoimmune Disease of the Ovary and Autoimmune Gastritis

  Materials
  • recipeBouins's fixative (see recipe)
  • 70% ethanol
  • Surgical instruments
  • Additional reagents and equipment for evaluation of autoimmune ovarian disease (unit 15.17, protocol 2), and paraffin embedding of tissue (unit 21.4)

Support Protocol 2: Detection of Serum Autoantibody to Ovarian and Gastric Antigens Using Indirect Immunofluorescence

  Materials
  • Freshly harvested adult mouse ovaries or fundus of mouse stomach
  • Phosphate‐buffered saline (PBS; appendix 2A)
  • 90% ethanol
  • Serum from autoimmune mice (see protocol 1)
  • Normal mouse serum as negative control for each experiment
  • Serum from a known positive control mouse with autoantibodies against target antigens (ovary, stomach) being tested, for each experiment
  • recipeImmunofluorescence (IF) buffer (see recipe)
  • Fluorescein‐conjugated goat or rabbit anti–mouse IgG (Jackson Immunoresearch Laboratories)
  • recipeElvanol in buffered glycerol (see recipe)
  • Coplin jars or staining dishes
  • Pap pen (Research Products International or Shandon/Lipshaw)
  • Fluorescence microscope
  • Additional reagents and equipment for immunohistochemistry of frozen tissue sections (see unit 21.4)
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Figures

Videos

Literature Cited

Literature Cited
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