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. Author manuscript; available in PMC: 2008 Feb 21.
Published in final edited form as: J Natl Cancer Inst. 2007 Feb 21;99(4):309–321. doi: 10.1093/jnci/djk053

Fig. 4.

Fig. 4

KISS1 secretion and lung metastasis by C8161.9 human melanoma cells. Cells were not transfected (P) or were transfected with empty vector (V), with FLAG-tagged KISS1 (KFM), or with FLAG-tagged KISS1 lacking the signal sequence (KFMΔSS). Numbers indicate KFM transfectant clones (17, 46, 57, 59, 66) and KFMΔSS transfectant clones (6, 7, 10, 16). Mix indicates uncloned transfectants for each KISS1 construct. A and D) Immunoblot analysis of conditioned medium from P, V, and KFM (A) or P, V, and KFMΔSS (D) cells using immunoprecipitation with anti-metastin polyclonal antiserum followed by immunoblotting with anti–FLAG M2 antibody. B and E) Relative quantification of KISS1 secretion levels by enzyme-linked immunosorbent assay of KFM (B) and KFMΔSS (E) clones. Removal of the putative signal sequence blocks secretion of KISS1 (E). KFM clone 59 is shown on panel E as a positive control for the assay. C and F) Lung colonization in mice injected intravenously with C8161.9 cells transfected with KFM (C) or KFMΔSS (F) was determined by counting surface lung foci. Statistical significance (*** P<.001) was calculated by comparison of experimental values (KFM versus KFMΔSS) with vector control (V) (Holm–Sidak method).