Submitted to: Experimental Biology
Publication Type: Abstract Only
Publication Acceptance Date: April 1, 1998
Publication Date: N/A
Technical Abstract:
A polyclonal antiserum was produced in rabbits immunized with a 15 amino acid peptide (QRVTGLDFIPGLHPV) derived from the reported sequence for porcine leptin. This peptide contained a core 8 amino acid sequence (GLDFIPGL) that is totally conserved for all reported leptins. Recombinant human, mouse and pig leptins were subjected to SDS-PAGE and electroblotted onto nitrocellulose. Western immunoblots were developed with an alkaline phosphatase second antibody system. The antiserum was highly specific for leptin which demonstrated a molecular weight of approximately 16 kDa. Sensitivity of the Western assay did not permit the direct detection of leptin in serum. Instead, plasma or serum samples from pig, sheep, cow, chicken and turkey blood were fractionated using FPLC and/or Sephacryl S-300 chromatography. Column fractions were collected, concentrated and analyzed by SDS-PAGE and Western blotting. Leptin was detected in a low molecular weight column fraction of chicken and turkey plasma, but not in pig, sheep or cow samples. The apparent molecular weight of avian leptin was approximately 13 kDa which is lower than the recombinant leptins. This could indicate possible differences between native and recombinant leptin proteins.
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