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The PVDF membrane was subsequently washed (5 min 5) with tris-buffered saline with tween (TBST) buffer and then incubated with the appropriate secondary antibody (Peroxidase-conjugated goat anti-rabbit IgG (H + L), at a dilution of 1:5000) in a 5% bovine serum albumin solution for 1 h at 25 C
If initial staining was insufficient, the sample was sectioned again to obtain a better visual field of cartilage structure
Weekly intervals resulted in an increase of tHcy concentration in plasma above 60 mol/l
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