Volume 116, Issue 3 , Pages 704-705, September 2005
IgE-binding peptides coupled to a commercial matrix as a diagnostic instrument for persistent cow's milk allergy
Article Outline
To the Editor:
Cow's milk allergy (CMA) is the most common cause of food allergy in the first years of life, affecting approximately 2.5% of children.1 The majority of children outgrow their CMA by 3 to 4 years of age; however, about 15% of children retain their allergy into the second decade.2, 3 The mechanisms behind this development of clinical tolerance to food proteins remain poorly understood.
In our previous studies, we characterized the sequential IgE-binding epitopes for the major cow's milk allergens.4, 5, 6, 7 Observed differences between younger children who are likely to outgrow their allergy and older patients with persistent cow's milk allergy suggested that epitope specificity of IgE antibody responses might predict the clinical outcome of CMA.4 Using SPOTs-membrane technology and sera from 10 patients with persistent and 10 patients with transient CMA, we identified informative IgE-binding epitopes that were not recognized by any of the patients with transient CMA but showed binding by most of the patients with persistent allergy.8 The current study was performed to confirm the relevance of these informative epitopes in a larger study population.
Therefore, 74 patients with challenge-proven CMA were included in the study. Twenty-nine patients subsequently outgrew their allergy, 15 of those at a median age of 3 years (range, 1-4 years; group C) and 14 at a median age of 8 years (range, 6-18 years; group B). The remaining 45 children followed until a median age of 8 years (range, 5-22 years; group A) retained their allergy, and were assumed to have persistent CMA.
Five peptides, 10 amino acids in length, representing the IgE-binding epitopes on αs1-casein, αs2-casein, and κ-casein, were synthesized and coupled to ImmunoCAP (Pharmacia Diagnostics, Uppsala, Sweden). These peptides were chosen from previously identified peptides because they were recognized solely by patients with persistent CMA.8 The decapeptides were coupled to the cyanogen bromide–activated cellulose solid phase matrix, ImmunoCAP, at optimal conditions: for αs1-casein, AAs 69-78 (EEIVPNSVEQ) and AAs 123-132 (MKEGIHAQQK); for αs2-casein, AAs 33-42 (ENLCSTFCKE) and AAs 171-180 (YQKFALPQYL); and for κ-casein, AAs 155-164 (SPPEINTVQV). Peptide-coupled ImmunoCAPs and reagents were kindly provided by Pharmacia Diagnostics. Levels of cow's milk and peptide-specific IgE were determined using the Pharmacia UniCAP system. Banked sera obtained at the time of clinical reactivity and stored at −80°C were used for analyses. Thirteen of the 74 patient sera used in the current study were also used in our previous study on informative epitopes.8
Milk peptide–specific IgE was significantly higher in patients with persistent CMA at a median age of 8 years (group A) compared with children who subsequently outgrew their allergy in the first few years of life at a median age of 3 years (group C) or patients who lost their allergy at a median age of 8 years (group B; P < .001 and P < .05, respectively; Fig 1). Specifically, only 2 of 15 patients in group C and 5 of 14 patients in group B recognized any of the selected peptides at the time of clinically relevant milk allergy, whereas 30 of 45 patients in group A did so. Median milk-specific IgE levels were also significantly different among the 3 patient groups (Fig 1). However, the range was from <1.0 to >100 kU/L in all 3 groups, resulting in an overlap not observed in this way for the milk peptide–specific IgE. Fig 2 shows the level of peptide-specific IgE for the individual peptides in all 3 groups of patients. The antibody pattern was heterogeneous in that the highest levels for the individual peptides were found in different patient sera.
Fig 1.
Milk and milk peptide–specific IgE levels in patients with persistent CMA (group A) compared with children with subsequent late (group B) or regular tolerance development (group C) at time of clinical reactivity.
Fig 2.
Peptide-specific IgE for individual peptides at time of clinically relevant allergy in 3 groups of patients: (A) with persistent CMA, and (B) with late and (C) with regular tolerance development. Letters highlight the patients with the highest levels for the individual peptides.
In summary, our current study confirmed that IgE antibodies to distinct allergenic epitopes of cow's milk proteins can be used as a marker of persistent CMA. The successful linkage of these peptides to a commercial matrix should enable the conduct of large prospective studies and simplify future diagnostic approaches.
References
- . Food allergy, part 1: immunopathogenesis and clinical disorders. J Allergy Clin Immunol. 1999;103:717–728
- . The natural history of food allergy. Pediatrics. 2003;111:1631–1637
- . Clinical course of cow's milk protein allergy/intolerance and atopic diseases in childhood. Pediatr Allergy Immunol. 2002;13(suppl 15):23–28
- . Identification of IgE- and IgG-binding epitopes on alpha(s1)-casein: differences in patients with persistent and transient cow's milk allergy. J Allergy Clin Immunol. 2001;107:379–383
- . Identification of IgE and IgG binding epitopes on beta- and kappa-casein in cow's milk allergic patients. Clin Exp Allergy. 2001;31:1256–1262
- . Identification of sequential IgE-binding epitopes on bovine alpha(s2)-casein in cow's milk allergic patients. Int Arch Allergy Immunol. 2002;129:93–96
- . IgE and IgG binding epitopes on alpha-lactalbumin and beta-lactoglobulin in cow's milk allergy. Int Arch Allergy Immunol. 2001;126:111–118
- . B-cell epitopes as a screening instrument for persistent cow's milk allergy. J Allergy Clin Immunol. 2002;110:293–297
Supported by a grant from the National Institutes of Health (AI-44236).Disclosure of potential conflict of interest: K. Beyer has received grants/research support from Danone. S. Ahlstedt and L. Venemalm are employed by Pharmacia Diagnostics. H. Sampson has a consultant arrangement with Seer, Inc, and has received grants/research support from the National Institutes of Health and Pharmacia Diagnostics (through Mount Sinai School of Medicine).
PII: S0091-6749(05)01288-1
doi:10.1016/j.jaci.2005.05.007
© 2005 American Academy of Allergy, Asthma and Immunology. Published by Elsevier Inc. All rights reserved.
Volume 116, Issue 3 , Pages 704-705, September 2005
