Development of selective antibodies against the human somatostatin receptor subtypes sst1-sst5.
Antisera selective for five somatostatin receptor subtypes, human sst1-sst5, were raised in rabbits. C-terminal parts of human sst1-sst5 receptors were expressed as fusion proteins with glutathione S-transferase. Fusion proteins were affinity-purified and used for raising polyclonal antibodies. In Western blot analysis, all five antisera were tested on preparations of mammalian cell lines transfected with human sst1-sst5, respectively. sst1 antiserum reacted with a broad band of 53-72 kDa. A band of 71-95 kDa was detected with the antiserum raised against sst2, 65-85 kDa with sst3 antiserum, 45 kDa with sst4 antiserum and 52-66 kDa with sst5 antiserum. No cross-reactivity could be detected to any of the other four somatostatin receptor subtypes. Enzymatical deglycosylation of the receptors revealed that sst1, sst2, sst5 and possibly sst3 in this system are subjected to N-linked glycosylation, whereas sst4 is not. Two of the antisera (sst2 and sst5) were used for immunohistochemical localization of the receptors. sst2 and sst5 antisera labeled neurons in e.g. the amygdaloid complex, hippocampus, fascia dentata and the neocortex in rat and monkey tissue. This is the first report on antisera against all five somatostatin receptor subtypes and the first immunohistochemical visualization of sst5 receptors in the mammalian brain.
Helboe
L
L
Department of Medical Anatomy, Panum Institute, University of Copenhagen, Denmark.
Møller
M
M
Nørregaard
L
L
Schiødt
M
M
Stidsen
C E
CE
D000679:Amygdala / Q000737:chemistry / Q000378:metabolism; D000818:Animals; D000906:Antibodies; D000918:Antibody Specificity; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007150:Immunohistochemistry; D008252:Macaca fascicularis; D008297:Male; D008970:Molecular Weight; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D016548:Prosencephalon / Q000166:cytology / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D017208:Rats, Wistar; D017481:Receptors, Somatostatin / Q000032:analysis* / Q000096:biosynthesis* / Q000145:classification; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis; D014162:Transfection
D000906:Antibodies; D017481:Receptors, Somatostatin; D011993:Recombinant Fusion Proteins