Studies on natural ST2 gene products in the human leukemic cell line UT-7 using monoclonal antihuman ST2 antibodies.
Eight species of murine monoclonal antibodies against human ST2 protein, which is highly similar in protein sequence to the interleukin 1 receptor, were produced. The fusion was carried out between the murine myeloma cell line PAI and murine lymph node or spleen cells from mice immunized with the recombinant ST2 protein produced in Escherichia coli. Characterization of these monoclonal antibodies by immunoblot analysis revealed that they all reacted with recombinant, N-glycosylated ST2 protein that was secreted from COS7 cells transiently transfected with a mammalian expression vector carrying ST2 cDNA. The recombinant N-glycosylated ST2 protein could be immunoprecipitated by 5 out of 6 species of the IgG class monoclonal antibodies. Furthermore, these antibodies were also able to detect, by immunofluorescence, the membrane-bound chimeric molecule possessing an extracellular portion of human ST2 and a transmembrane and cytoplasmic portion of murine receptor type ST2L expressed on COS7 cells, indicating that these monoclonal antibodies were useful for detecting the natural membrane-bound ST2 in human cells. Combining immunoprecipitation and immunofluorescence with the aid of these monoclonal antibodies, together with the reverse transcriptase-polymerase chain reaction method, the human leukemic cell line UT-7 was demonstrated to express human ST2 mRNA and protein. The identification of the ST2 gene product in UT-7 cells may help investigators elucidate the function of the human ST2 gene.
Yoshida
K
K
Department of Applied Biological Science, Faculty of Science and Technology, Science University of Tokyo, Chiba, Japan.
Arai
T
T
Yokota
T
T
Komatsu
N
N
Miura
Y
Y
Yanagisawa
K
K
Tetsuka
T
T
Tominaga
S
S
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis; D001483:Base Sequence; D005260:Female; D005822:Genetic Vectors / Q000276:immunology; D006801:Humans; D000072179:Interleukin-1 Receptor-Like 1 Protein; D007938:Leukemia / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D008565:Membrane Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011506:Proteins / Q000235:genetics* / Q000276:immunology*; D012333:RNA, Messenger / Q000096:biosynthesis; D011956:Receptors, Cell Surface; D018123:Receptors, Interleukin; D011994:Recombinant Proteins / Q000276:immunology; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; C079241:IL1RL1 protein, human; C486189:Il1rl1 protein, mouse; D000072179:Interleukin-1 Receptor-Like 1 Protein; D008565:Membrane Proteins; D011506:Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D018123:Receptors, Interleukin; D011994:Recombinant Proteins