Diagnosis of Food Allergy by Combined Detection of Short Circuit Current Changes and Histamine Release Induced by Antigen Application during Mucosa-Oxygenation in Human Intestinal Biopsies

Abstract

Diagnosis of food allergy is hampered by the poor sensitivity and specificity of RAST and skin prick tests. Double-blind placebo-controlled food challenge (DBPCFC), the golden standard, is time and cost consuming. We performed ex vivo tests with intestinal biopsies of two patients with food allergies diagnosed by DBPCFC, and with biopsies of one patient with no known food allergies. Biopsies were mounted in Ussing chambers, serosal surface area was 1.1mm2, both compartments contained 2ml oxygenated Ringer’s solution at 37o C. Short circuit current (Isc) responses to serosally applied food protein extracts (10–250µg/ml) from pork, soybean, wheat, barley, rye, maize and ovalbumin and β-lactoglobulin were analyzed. Serosal and mucosal samples of 100 m l were taken before antigen application and at 2.5, 5.0 and 7.5 minutes thereafter, and were analyzed for histamine content by ELISA. Isc-changes induced by 65 antigen application tests in 24 biopsies ranged from 0–17µA/cm2 and appeared within 1 to 5 minutes. Histamine concentration increments peaked within 5 minutes after antigen application and ranged from 0–0.89 ng/ml (serosal) and from 0–0.49 ng/ml (mucosal). Serosal and mucosal appearance of histamine after antigen application correlated significantly (r=0.68, p<0.0001), and Isc-changes after antigen application correlated significantly to serosal and also to mucosal histamine appearance (r=0.44, p=0.0003 and r=0.29, p=0.02). Isc-changes and histamine release (5.50±0.67µA/cm2, respectively serosal: 0.15±0.04 ng/ml, mucosal: 0.13±0.03 ng/ml; n=30) induced by DBPCFC-positive antigens were significantly higher (p<0.0005) than those by DBPCFC-negative antigens (0.65±0.31µA/cm2, respectively serosal: 0.01±0.01 ng/ml, mucosal: 0.02±0.01 ng/ml; n=17).Detectable Isc-changes and histamine concentration increments were observed in antral, jejunal, ileal, colonic and rectal biopsies after application of relevant food antigens proven positive in the DBPCFC. These results indicate that the combined detection of Isc-changes and of histamine release after application of antigens to living endoscopical biopsies ex vivo may provide a new, sensitive and specific tool in the diagnosis of gastrointestinal food allergy.