Anti-MMP13 antibody (ab39012)

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.	-Rabbit (1:4000) -To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for 5 min. Lysates can be aliquoted and stored at -20°C for future use.

Upstream tips
-These buffers may be stored at 4°C for several weeks or aliquoted and stored at -20°C for up to a year.
Protocol tips
-Rabbit (1:4000) -To reduce and denature your samples, boil each cell lysate in sample buffer at 100°C for 5 min. Lysates can be aliquoted and stored at -20°C for future use.

Publication protocol

The NP cells from each culture dish were collected and lysed in RIPA buffer. The lysates were centrifuged, and the protein concentrations were determined by the BCA protein assay. In each group, equivalent amounts of protein (50 μg) were used for electrophoresis. After electrophoresis, the proteins were electrotransferred to 0.45 μm-pore-diameter polyvinylidene difluoride (PVDF) membranes (Invitrogen). After being blocked in Tris-buffered saline containing Tween-20 with 5% milk powder, the membranes were immunolabeled with specific antibodies for PN-1 (1:1500), FN N-terminal (1:5000), P38 (1:800), P-P38 (1:1000), ERK (1:5000), P-ERK (1:800), P65(1:2000), P-P65 (1:800), MMP3(1:1000), MMP9(1:1000), MMP13 (1:4000), ADAMTS4 (1:1000), ADAMTS5 (1:80), Aggrecan (1:100), or COL2 (1:8000) overnight at 4 °C with gentle shaking. After washing, the membranes were incubated for 2 h at 4 °C with a 1:2000 dilution of HRP-conjugated secondary antibody in antibody diluents (Boster). Finally, the ECL Plus western blotting system was used, and immunoreactive bands were quantified using ImageQuant LAS 400 software (GE Healthcare Life Sciences) and calculated by normalization to the reference bands of β-actin or Lamin B.

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Manufacturer protocol

Download the product protocol from Abcam for Anti-MMP13 antibody (ab39012) below.

Download manufacturer protocol

Videos

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