Immunohistochemical detection of somatostatin sst2a receptors in the lymphatic, smooth muscular, and peripheral nervous systems of the human gastrointestinal tract: Facts and artifacts

The cellular distribution of the somatostatin sst2A receptor protein was in vestigated in the lymphatic, smooth muscular, and nervous components of the human gastrointestinal tract using subtype-specific antibody R2-88 for imm unohistochemical staining of cryostat and formalin-fixed, paraffin-embedded tissue sections. Germinal centers of intestinal lymphatic follicles were i mmunostained, exhibiting a predominantly plasma membrane localization of th e receptor. Similarly, nerve fibers and cells in the submucosal and myenter ic plexus were stained for sst2A. Antibody preabsorption with 100 nmol/L an tigen peptide abolished staining in all of these tissues, and immunohistoch emical staining correlated with the labeling observed after receptor autora diography using the sst2-preferring radioligand I-125-[Tyr(3)]octreotide. C ytoplasmic immunostaining was detected in gastrointestinal smooth muscle ce lls and was inhibited by antibody preabsorption with antigen peptide. Howev er, I-125-[Tyr(3)]octreotide autoradiography was negative, and Western blot s showed no band at the usual 70-90 kDa location for sst2A. Instead, a band was observed at 205 kDa. This band comigrated with the rabbit myosin stand ard, which was also stained with R2-88, although antibody sensitivity for m yosin was less than 0.002% of that for the sst2A receptor. Rigorous compute r-based sequence analysis demonstrated the peptide sequence chosen for anti body production was unique. Moreover, standard sequence alignment protocols were unable to identify the sequences in myosin responsible for the observ ed reactivity with the R2-88 antiserum. The observed cross-reactivity empha sizes the need for extensive controls to prove the specificity of immunosta ining for such low abundance proteins as receptors even when the peptide se quence chosen for antibody production is unique. This study demonstrates fo r the first time the presence of specific sst2A receptor protein by immunoh istochemistry in the human gastrointestinal lymphatic and nervous component s, but not in gastrointestinal circular and longitudinal smooth muscle.