Quantifying TGFβ Release from LTBP1 Matrix

TGFβ release was determined as described^{34 (link)}. Briefly, CHO LTBP1 cell were seeded (5.0 × 10⁴ cells per well) for 48 h in a 96 well plate in triplicates to deposit an LTBP1-TGFβ rich matrix. Cells were then detached with PBS/15 mM EDTA, the remaining LTBP1-TGFβ rich matrix washed twice with PBS and incubated with keratinocytes (2.0 × 10⁴ cells per well) and tMLEC cells (1.5 × 10⁴ cells per well) in a 100 μl final volume either in the absence or presence of α_vβ₆ integrin blocking antibody (10D5; 20 μg ml⁻¹; Chemicon) or TGFβ neutralizing antibody (1D11; 15 μg ml⁻¹; BD Biosciences). The amount of released TGFβ was measured after 16–24 h using Bright Glo luciferase Kit (Promega).