Fig. 6.
Fig. 6. (top) Western immunoblots of cell lysates isolated from H-FT–transfected clones. Depicted are the ECL exposures of the trans-blots of SDS-PAGE of the respective samples (same protein load on gels) derived from the different clones using the following antibodies: anti-PgP mouse monoclonal, rabbit antimouse H-FT and anti–L-FT antibodies, rabbit antiactin and the respective goat antimouse or antirabbit IgG conjugated to HRP. The samples were run in parallel on separate gels, but the exposure times differed for the different antibodies used. The numbers above the bands represent the values of the densitometry tracings using the intensity of cl-16 for normalization (center). Correlation between H-FT and PgP levels of expression in MEL clones. The densitometry tracings of the immunoblots shown on the top and others (not shown) were normalized to the values obtained in cl-16 for both H-FT and MDR. The denstity values of each pair (n = 3) were from parallel SDS-PAGE runs originating from the same cell samples normalized to that of actin. The mean OD values are given as symbols and the ± SE as bars both PgP and H-FT, respectively (bottom). RT-PCR of H-FT, MDR1, and L7 ribosomal mRNAs of a high H-FT expresser and the wild-type untransfected clone. The propidium iodide stains are of the various mRNA samples analyzed by RT-PCR as described in Methods. For the relatively abundant H-FT message the system saturated at cycle 18, whereas for MDR1a and L7 it had to be run at higher cycle number. MDR1a was essentially undetected in the wild type (wt).

(top) Western immunoblots of cell lysates isolated from H-FT–transfected clones. Depicted are the ECL exposures of the trans-blots of SDS-PAGE of the respective samples (same protein load on gels) derived from the different clones using the following antibodies: anti-PgP mouse monoclonal, rabbit antimouse H-FT and anti–L-FT antibodies, rabbit antiactin and the respective goat antimouse or antirabbit IgG conjugated to HRP. The samples were run in parallel on separate gels, but the exposure times differed for the different antibodies used. The numbers above the bands represent the values of the densitometry tracings using the intensity of cl-16 for normalization (center). Correlation between H-FT and PgP levels of expression in MEL clones. The densitometry tracings of the immunoblots shown on the top and others (not shown) were normalized to the values obtained in cl-16 for both H-FT and MDR. The denstity values of each pair (n = 3) were from parallel SDS-PAGE runs originating from the same cell samples normalized to that of actin. The mean OD values are given as symbols and the ± SE as bars both PgP and H-FT, respectively (bottom). RT-PCR of H-FT, MDR1, and L7 ribosomal mRNAs of a high H-FT expresser and the wild-type untransfected clone. The propidium iodide stains are of the various mRNA samples analyzed by RT-PCR as described in Methods. For the relatively abundant H-FT message the system saturated at cycle 18, whereas for MDR1a and L7 it had to be run at higher cycle number. MDR1a was essentially undetected in the wild type (wt).

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