Production and isolation of guinea pig IgE antibody.
Immunoglobulins of the IgE and IgG classes have been causally associated with hypersensitivity reactions in man and in numerous animal species including mice, rats and guinea pigs. The use of the guinea pig as an animal model for both pulmonary and dermal hypersensitivity reactions, and the recent recognition of the importance of IgE antibodies in both early- and late-onset hypersensitivity responses, has heightened interest in production, separation, and isolation of this immunoglobulin class from the guinea pig. IgE antibodies were produced by treatment of strain 13 guinea pigs with cyclophosphamide followed by injection with S. aureus enterotoxin. Serum was obtained and the globulin fraction isolated by addition of caprylic acid then ammonium sulfate. Immunoglobulins were separated into classes using fast protein liquid chromatography (FPLC) employing a Mono Q column and a linear gradient of 0.01-0.3 M Na,K phosphate buffer, pH 7.5 (buffer B). IgG eluted in two major peaks. IgG2 was not retained on the column and emerged with the starting buffer; IgG1 was eluted with 15-20% buffer B. IgE, detected as heat labile homocytotropic antibody, was found in the fraction eluting with 30-35% buffer B. The elution profile of the guinea pig immunoglobulins was predicted from the pattern obtained with immunoglobulin classes from other species. This chromatographic procedure enabled rapid isolation of immunoglobulin classes from guinea pig sera and effectively separated IgG1 from IgE, the two classes associated with hypersensitivity reactions.
Karol
M
M
Department of Environmental and Occupational Health, Graduate School of Public Health, University of Pittsburgh, PA 15261.
Jin
R Z
RZ
Bennedsen
M
M
Vaughan
F
F
D000818:Animals; D000906:Antibodies / Q000302:isolation & purification*; D000917:Antibody Formation; D002845:Chromatography; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006168:Guinea Pigs; D007073:Immunoglobulin E / Q000096:biosynthesis / Q000276:immunology / Q000302:isolation & purification*; D010323:Passive Cutaneous Anaphylaxis
D000906:Antibodies; D007073:Immunoglobulin E