Methods:
Methods related to the constructs, purification, biophysical characterisation, NMR, and microarray statistical analysis are located in the supplemental information.

Patient sera

Sera from 40 individuals with walnut (12 patients), peanut (12 patients), and peanut/walnut allergy (16 patients) were collected in accordance with rules and regulations of the institutional review board of their respective institutions (Tulane University Biomedical IRB 09-00231, REF #: 140613; University of California at Davis IRB protocol No. 200210194-6) and in accordance with U.S. federal policy for the protection of human subjects (Table 1). Patients were over the age of 18 and had experienced recurrent severe, systemic allergic reactions to peanut, walnut or both. These patients were included in this study based on their convincing clinical history of food allergy and did not undergo food challenge due to the potential severity of reactions.

Microarray

The entire amino acid sequences of Ara h 1 and Jug r 2 were printed onto microarray slides (JPT Peptide Technologies GmbH, Berlin, Germany) as sequential overlapping 15 amino acid spots, offset by 5 amino acids. Slides were placed into a HS400 Pro (Tecan, San Jose, CA), blocked in filtered SuperBlock TBS (Thermo Fisher Scientific, Waltham, MA) for 30 minutes at room temperature under agitation and then washed with Tris-buffered saline containing 0.5% Tween-20 (TBST) (Bio-Rad, Hercules, California). After centrifugation, 200µL of each patient’s undiluted sera was injected into individual chambers containing microarray slides and incubated at 4°C for 16 hours with agitation. Microarray slides were then washed and injected with 170 µL of mouse α-human IgE (Life Technologies, Grand Island, NY) diluted into filtered Superblock at a dilution of 1:5000 for 30 minutes at room temperature. The slides were washed and dried before scanning on a GenePix-4000B (Molecular Devices, San Jose, CA). IgE binding was measured by Cy3 green fluorescence at 532nm. The data was analyzed by GenePix Pro 7.2 software.
Statistics
Statistical analyses were performed using R (version 3.6.3). Modified z values were calculated from the microarray median signal-to-noise ratios (SNRs) and used to determine the percent of leader sequence peptides bound by IgE for each allergy group. Median SNR values were converted into modified z values, calculated using median and median absolute deviation (MAD) of a patient’s SNRs to an entire leader sequence or leader sequence fragment. To approximate standard deviation, MAD was calculated using the constant of 1.4826. We defined a positive binding event as a z value >= 3. Using the fisher.testfunction, Fisher’s exact tests were used to compare the proportion of positive binding events out of the total possible binding events for a particular fragment among allergy groups. When Fisher’s tests indicated a significant difference between one or more allergy groups (p-value < 0.05), pairwise post hoc comparisons were made to determine which allergy groups differed from one another. We defined major epitopes as peptides positively bound by 50% or more or patients within an allergy group. Average z values which directly relate to IgE binding intensity are presented as low (3 – 6), medium (6.1 – 9) and high (>9).  See supplemental methods for more details.