No abstract

SUMMARY Serotonin (5-hydroxytryptamine, 5-HT) mediates many functions of the central and peripheral nervous systems by its interaction with specific neuronal and glial receptors. Fourteen serotonin receptors belonging to seven families have been identified through physiological, pharmacological, and molecular cloning studies. Monoclonal antibodies (MAbs) specific for each of these receptor subtypes are needed to characterize their expression, distribution, and function in embryonic, adult, and pathological tissues. In this article we report the development and characterization of MAbs specific to the serotonin 5-HT 2A receptor. To generate MAbs against 5-HT 2A R, mice were immunized with the N-terminal domain of the receptor. The antigens were produced as glutathionine S-transferase (GST) fusion proteins in insect cells using a Baculovirus expression system. The hybridomas were initially screened by ELISA against the GST-5-HT 2A R recombinant proteins and subsequently against GST control proteins to eliminate clones with unwanted reactivity. They were further tested by Western blotting against recombinant GST-5-HT 2A R, rat and human brain lysate, and lysate from cell lines transfected with 5-HT 2A R cDNA. One of the MAbs G186-1117, which recognizes a portion of the 5-HT 2A R N-terminus, was selected for further characterization. G186-1117 reacted with a band of molecular size 55 kD corresponding to the predicted size of 5-HT 2A R in lysates from rat brain and a 5-HT 2A R-transfected cell line. Its specificity was further confirmed by adsorption of immunoreactivity with recombinant 5-HT 2A R but not with recombinant 5-HT 2B R and 5-HT 2C R. Rat brain sections and Schwann cell cultures were immunohistochemically labeled with this MAb. G186-1117 showed differential staining in various regions of the rat brain, varying from regions with no staining to regions of intense reactivity. In particular, staining of cell bodies and dendrites of the pyramidal neurons in the cortex was observed, which is in agreement with observations of electrophysiological studies. (J Histochem Cytochem 46:811-824, 1998)

No abstract

No abstract

Nonrandom chromosomal abnormalities are found in most human malignancies, particularly leukemias and lymphomas. A characteristic t(1; 19) (q23; p13.3) chromosomal translocation is detected in 5% of childhood acute lymphoblastic leukemia (ALL) cases. This translocation results in the formation of a fusion gene, which leads to the expression of an oncogenic E2A/pbx1 protein. Breakpoints in the E2A gene almost invariably occur within a single intron, and the identical portion of PBX1 is joined consistently to exon 13 of E2A in fusion mRNA. In this article, we report the development of monoclonal antibodies against E2A/pbx1 fusion protein using a specific peptide that corresponds to the junction region of the protein. The obtained antibodies recognize specifically the chimeric E2A/pbx1 fusion protein and lack cross-reactivities with E2A and pbx1. Immunohistochemical staining and flow cytometric studies show that these antibodies can distinguish t(1; 19)-positive from t(1; 19)-negative leukemic cells. These results indicate that the obtained E2A/pbx1-specific monoclonal antibodies might prove to be valuable diagnostic reagents and important tools for elucidating the mechanisms involved in oncogenesis and progression of t(1; 19)-positive childhood ALL.