A ten-year retrospective analysis of the distribution, use and phenotypic characteristics of the LAD2 human mast cell line.

BACKGROUND: In 2003, this laboratory published an account of the human mast cell line LAD2 (Laboratory of Allergic Diseases 2) that expressed FcεRI, responded to recombinant human stem cell factor (rhSCF) and resembled CD34+-derived human mast cells. LAD2 cells have now been distributed worldwide. To study the impact of this transfer, we analyzed the number of investigators receiving LAD2 cells and resulting publications. METHODS: Records maintained in our laboratory, the Technology Transfer and Intellectual Property Office and Office of Technology Transfer, were reviewed for material transfer agreements (MTAs) and licensing agreements (LAs). Journals and impact factors were obtained from PubMed.gov by cross-referencing LAD2 and human mast cells from 2003 through November 2013. RESULTS: Over 300 MTAs and 40 LAs were approved. LAD2 cells were shipped to over 30 countries. More than 80 papers have been published in journals with impact factors from 1.31 to 13.21. Intended uses include the study of receptors, degranulation, and cell signaling. LAD2 cells continue to express described markers and have consistent FcεR1-mediated degranulation. CONCLUSIONS: Success of the LAD2 line reflects the demand for a human mast cell line in research, the uniqueness of this cell line, and that it continues to exhibit minimal variation from its original description. We hope that the awareness of the impact of this cell line on mast cell research will encourage others to develop and distribute other similar cell lines with additional characteristics so as to address the limitations of depending on the study of cultured human mast cells from tissues.

Energy transfer ligands of the GluR2 ligand binding core.

Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that mediate excitatory signaling in the central nervous system. When a ligand binds to the extracellular domain of iGluRs, local conformational changes ensue and this motion is translated to the transmembrane domain, inducing channel opening. We have used an isolated ligand binding domain, GluR2-S1S2J (GluR2), as a model system to study the protein-ligand complex by steady-state and time-resolved intrinsic tryptophan fluorescence measurements. Specifically, we determined that the widely used and structurally characterized antagonist, 6,7-dinitroquinoxaline-2,3-dione (DNQX), acts as an efficient fluorescence energy transfer (FET) acceptor for Trp. Consistent with crystallographic data, our results indicate that the four native tryptophans are within Forster's radius (R(o) approximately 33 A) of the bound ligand. Additionally, we demonstrate the broader value of this technique by identifying an original FET ligand, 3-nitrotyrosine (3NY), for GluR2 (R(o) approximately 24 A; apparent dissociation constant K(d) approximately 170 microM). Estimated average donor-acceptor (Trp-ligand) distances extracted from tryptophan excited-state decays are similar for both ligands (approximately 24 A), suggesting that 3NY binds in the structurally characterized ligand binding cleft. Moreover, an alternative competition assay utilizing Trp --> DNQX quenching for detection of ligand binding in GluR2 is described.