MMP requirement for spheroid cell penetration into 3D matrices is matrix specific. Ovarian cancer cell spheroids, generated by hanging drop culture, were embedded in polymerized matrices of A, Matrigel (undiluted, ~11 mg/ml) B, pepsin-extracted collagen I, (Vitrogen) and C, acid-extracted collagen I (2.1 mg/ml). Images reveal the spheroid edge, where cell penetration of matrices was apparent. An entire spheroid is shown in the lower inset. Cell penetration of Matrigel and pepsin-extracted collagen I (Vitrogen) occured despite MMP blockade with GM6001. In contrast, invasion of acid-extracted collagen 1 was prevented by MMP inhibition. Representative results are shown using OVCA429 spheroids at 60, 36, and 44 hrs for Matrigel, Vitrogen and acid-extracted collagen I, respectively.