Figure 4

Immunoblot Analysis of Cell and Tissue Lysates: Lysates were made of normal human mammaryepithelia and tumor cell lines. Cell lysates were made, and soluble proteinswere loaded at a final concentration of 1 mg/ml. Samples were separated on 10%SDS-PAGE in a Mini-Protean II apparatus (25μg/sample), transferred tonitrocellulose, and probed with the anti-Mammastatin monoclonal antibody,7G6.6, or the IgM control Antibody 38C13 developed colorometricaly withNBT/BCIP. A. (Normal Mammary Cells) Lane 1) NHMC-17, 2) NHMC-16, 3) NHMC-15, 4)NHMC-14, 5) NHMC-6, Repeated many times with similar results. B. (CarcinomaCell Lines) Lane 1) SK BR-3, 2) Du 4475, 3) MDA MB-361, 4) ZR-75-30, 5) BT-549,6) MCF-7, 7) Sum 102, 8) Sum 52, 9) Sum 44, 10) NHMC-21, C. (Breast TumorTissue) 1) DW tumor lysate, 2) IR tumor lysate, 3) EW tumor lysate, 4) PH tumorlysate, 5) SE tumor lysate, 6) AW tumor lysate, 7) EJ tumor lysate; 8) SPnormal tissue (from cancer patient), 9) NHMC-12 organoid. Repeated three timeswith similar results, D. (Primary Culture of Breast Tumor Tissue) Lane 1)CHMC-42, 3 days, 2) CHMC-42 tissue, 3) CHMC-39, two days, 4) CHMC-39 tissue, 5)BHMC-40, three days, 6) BHMC-40 tissue, 7) ZR-75-1 carcinoma, 8) MDA 435carcinoma, 9) MDA 231 carcinoma, 10) NHMC-20 primary culture.