Development of an Enzyme-Linked Immunosorbent Assay for Quantitative Determination of TSLP in Cell Culture Medium and Human Serum (38.20)

Abstract

Abstract Human thymic stromal lymphopoietin (TSLP) is a 24 kDa monomeric glycoprotein produced primarily by non-hematopoietic cells such as fibroblasts, epithelium, and stromal cells. TSLP induces myeloid cells to release T cell chemoattactents while enhancing maturation of CD11c+ dendritic cells that in turn promote a Th2 immune phenotype. We have developed a sandwich ELISA to quantitatively measure human TSLP concentrations in culture media and human serum. The capture and detection antibodies consist of antigen affinity-purified goat anti-human TSLP polyclonal antibodies generated from recombinant human TSLP as the immunogen. The assay has a linear detection range from 31 to 250 pg/ml, and a threshold of detection at approximately 2 pg/ml. Serum spike recovery averaged 92.6% for four human serums (2 male, 2 female) for spike concentrations from 31.25pg to 500pg/ml inclusive. Assay robustness was further demonstrated by acceptable linearity of sample dilutions, intra-assay and inter-assay precisions. When primary normal human bronchial epithelial cells were stimulated with dsRNA and/or IL-4, TSLP concentration in the culture supernatant of the double stimulated cells increased significantly (53pg/ml, sd 3.37) compared with that from dsRNA stimulated cells (22pg/ml, sd 1.5, p=0.001). Unstimulated cell media had no detectable TSLP. This assay will provide a valuable resource for the study of role of TSLP in normal and pathophysiological conditions.