Assessment of three human FcepsilonRI-transfected RBL cell-lines for identifying IgE induced degranulation utilizing peanut-allergic patient sera and peanut protein extract.
Specific IgE sera screening studies are employed to investigate protein cross-reactivity. Such nonfunctional immunochemical methods cannot measure the biological activity of proteins. Therefore, an assay using RBL cells transfected with human FcepsilonRI was developed. Our objective was to evaluate the degranulation of three cell-lines expressing either the alpha-(RBL-hEI(a)-2B12 and RBL-30/25cells) or alpha-, beta-, and gamma-subunits (RBL SX-38) of the human FcepsilonRI by beta-hexosaminidase release. Purified human IgE and serum-derived polyclonal IgE from peanut-allergic subjects following challenge with anti-IgE or peanut protein extract, respectively, were utilized. Robust degranulation was induced in all three: RBL-30/25 (84%), -hEI(a)-2B12 (54%), SX-38 (94%), respectively, using purified IgE+anti-human IgE. Good release (18%, 40-45%, and 65%, respectively) occurred for one peanut-allergic subject+peanut extract with all cell-lines. With serum from three other peanut-allergic subjects, no beta-hexosaminidase release occurred with RBL-hEI(a)-2B12 cells+peanut extract, while only serum from one subject induced good degranulation, 30% and 60%, respectively, with RBL-30/25 and RBL SX 38 cells. Consistent degranulation with a potent food allergen (peanuts) was not observed. The assay's utility in safety assessment, predictive value and reproducibility for evaluating the cross-reactivity of proteins with allergens needs further investigation with additional proteins and well-characterized sera.
Ladics
G S
GS
DuPont Company, Building 353/334, Route 141 and Henry Clay Road, Wilmington, DE 19880, USA. Gregory.s.ladics@usa.dupont.com
van Bilsen
J H M
JH
Brouwer
H M H
HM
Vogel
L
L
Vieths
S
S
Knippels
L M J
LM
D000485:Allergens / Q000097:blood / Q000276:immunology*; D000818:Animals; D010367:Arachis / Q000737:chemistry / Q000276:immunology*; D015550:Cell Degranulation / Q000187:drug effects / Q000276:immunology*; D045744:Cell Line, Tumor; D004306:Dose-Response Relationship, Immunologic; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood / Q000235:genetics / Q000276:immunology*; D008407:Mast Cells / Q000166:cytology / Q000187:drug effects / Q000276:immunology*; D021183:Peanut Hypersensitivity / Q000097:blood / Q000276:immunology*; D010936:Plant Extracts / Q000276:immunology / Q000633:toxicity; D051381:Rats; D017455:Receptors, IgE / Q000235:genetics / Q000378:metabolism; D014162:Transfection / Q000379:methods; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
D000485:Allergens; D010936:Plant Extracts; D017455:Receptors, IgE; D007073:Immunoglobulin E; D001619:beta-N-Acetylhexosaminidases