Development of selective antibodies against the human somatostatin receptor subtypes sst 1–sst 5

Abstract

Antisera selective for five somatostatin receptor subtypes, human sst 1–sst 5, were raised in rabbits. C-terminal parts of human sst 1–sst 5 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 sst 1–sst 5, respectively. sst 1 antiserum reacted with a broad band of 53–72 kDa. A band of 71–95 kDa was detected with the antiserum raised against sst 2, 65–85 kDa with sst 3 antiserum, 45 kDa with sst 4 antiserum and 52–66 kDa with sst 5 antiserum. No cross-reactivity could be detected to any of the other four somatostatin receptor subtypes. Enzymatical deglycosylation of the receptors revealed that sst 1, sst 2, sst 5 and possibly sst 3 in this system are subjected to N-linked glycosylation, whereas sst 4 is not. Two of the antisera (sst 2 and sst 5) were used for immunohistochemical localization of the receptors. sst 2 and sst 5 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 sst 5 receptors in the mammalian brain.