Generation and characterization of subtype-specific monoclonal antibodies to K+ channel alpha- and beta-subunit polypeptides.
Molecular characterization of mammalian voltage-sensitive K+ channel genes and their expression became possible with the cloning of the Shaker locus of Drosophila. However, analysis of the expression patterns and subunit composition of native K+ channel protein complexes requires immunological probes specific for the individual K+ channel gene products expressed in excitable tissue. Here, we describe the generation and characterization of monoclonal antibodies (mAbs) against eight distinct mammalian K+ channel polypeptides; the Kv1.1, Kv1.2, Kv1.4, Kv1.5 and Kv1.6 Shaker-related alpha-subunits, the Kv2.1 Shab-related alpha-subunit, and the Kv beta 1 and Kv beta 2 beta-subunits. We characterized the subtype-specificity of these mAbs against native K+ channels in mammalian brain and against recombinant K+ channels expressed in transfected mammalian cells. In addition, we used these mAbs to investigate the cellular and subcellular distribution of the corresponding polypeptides in rat cerebral cortex, as well as their expression levels across brain regions.
Bekele-Arcuri
Z
Z
Department of Biochemistry and Cell Biology, State University of New York, Stony Brook 11794, USA.
Matos
M F
MF
Manganas
L
L
Strassle
B W
BW
Monaghan
M M
MM
Rhodes
K J
KJ
Trimmer
J S
JS
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D000918:Antibody Specificity; D001923:Brain Chemistry; D019556:COS Cells; D002540:Cerebral Cortex / Q000737:chemistry; D051668:Delayed Rectifier Potassium Channels; D019084:Fluorescent Antibody Technique, Indirect; D015151:Immunoblotting; D051663:Kv1.2 Potassium Channel; D051665:Kv1.4 Potassium Channel; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology*; D015221:Potassium Channels / Q000032:analysis / Q000276:immunology*; D024642:Potassium Channels, Voltage-Gated; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000032:analysis / Q000276:immunology; D051669:Shab Potassium Channels; D014162:Transfection
D000911:Antibodies, Monoclonal; D051668:Delayed Rectifier Potassium Channels; C496970:Kcna2 protein, mouse; C496972:Kcna2 protein, rat; C496978:Kcna4 protein, mouse; C496979:Kcna4 protein, rat; C496990:Kcnb1 protein, mouse; C496991:Kcnb1 protein, rat; D051663:Kv1.2 Potassium Channel; D051665:Kv1.4 Potassium Channel; D010455:Peptides; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011994:Recombinant Proteins; D051669:Shab Potassium Channels