BioMed Research International / 2010 / Article / Fig 4

Research Article

The Collagen V Homotrimer Production Is Unexpectedly Favored over the Heterotrimer in Recombinant Expression Systems

Figure 4

Characterization of the recombinant procollagen V molecular forms produced in HEK-293 transfected cells. (a) Western blot analysis of the supernatant of transfected cell media dialysed against Tris-HCl 50 mM, pH 8.6, digested (lane 2) or not (lane1) with pepsin. Transfected cell media were centrifuged and the supernatant was applied to a DEAE column. (b) Separation of the different collagen V molecular species from transfected cell media by cation exchange chromatography. Western blot analysis of the loaded sample (lane 1) and of the fractions eluted at 0.16 M (lane 2), 0.24 M (lane 3), 0.37 M (lane 4), and 0.42 M (lane 5) NaCl. Membranes were double probed with mAb 18G5 followed by alkaline phosphatase antimouse secondary antibodies and with pAb CollV followed by peroxydase antirabbit secondary antibodies. Left, molecular mass standards expressed in kDa.

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