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Quantitative Enzyme-linked Immunosorbent Assay (ELISA) to Measure Serum Levels of Murine Anti-histone Antibodies
Published: Jun 20, 2012 DOI: 10.21769/BioProtoc.228 Views: 12057
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Abstract
Autoimmune disease is characterized with the break of tolerance against self antigens. Anti-histone antibodies can be found in the majority of patients with system lupus erythematosus (SLE) and a number of lupus-prone mouse strains. This protocol describes a reliable method to determine the relative serum titers of anti-histone antibodies in these mice.
Keywords: ELISA
Mouse
Lupus
Autoimmune
Anti-histone antibody
Materials and Reagents
- Histone from calf thymus (Roche Diagnostics, catalog number: 10223565001 )
- Phosphate buffered saline (PBS)
- Tween 20
- Na2HPO4 (anhydrous)
- NaH2PO4 (anhydrous)
- NaCl
- Fetal bovine serum (FBS) (Hyclone)
- Bovine serum albumin (BSA)
- Horseradish peroxidase (HRP) conjugated goat anti-mouse isotype specific antibodies [Southern Biotech, catalog number: 1040-05 (IgA); 1030-05 (IgG); 1021-05 (IgM)]
- ABTS Peroxidase Substrate Solution A and B (Kirkegaard & Perry Laboratories, catalog number: 50-62-01 )
- ABTS Peroxidase Stop Solution (Kirkegaard & Perry Laboratories, catalog number: 50-85-01 )
- 10x PBS-Tween 20 (see Recipes)
- Blocking solution (see Recipes)
Equipment
- Standard bench-top centrifuge
- Falcon 96-well ELISA plates (BD Biosciences, catalog number: 35-3915 )
- ELISA reader
- Parafilm
Procedure
Category
Immunology > Antibody analysis > Antibody detection
Biochemistry > Protein > Immunodetection
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