Autophagy Inhibitor, 3-MA

Protocol tips

Upstream tips	Protocol tips	Downstream tips
	Use 1mM concentration of 3-MA for in vitro treatment of cells

Protocol tips
Use 1mM concentration of 3-MA for in vitro treatment of cells

Publication protocol

The cells (1.5 × 106) were washed twice in ice cold PBS, re-suspended in 200 µl ice cold solubilizing buffer (300 mM NaCl, 50 mM Tris-Cl (pH 7.6), 0.5% Triton X-100, 2 mM phenylmethylsulfonyl fluoride (PMSF), 2 µl/ml aprotinin, and 2 µl/ml leupeptin) and incubated at 4℃ for 1 h. The lysates were centrifuged at 13,200 rpm for 30 min at 4℃. Protein concentrations of cell lysates were determined using a Bradford protein assay (Bio-Rad, Richmond, CA, USA), and 20 µg of protein was resolved using 10% sodium dodecyl sulfate (SDS)/polyacrylamide gel electrophoresis (PAGE) gel. The gel was transferred to polyvinylidene fluoride (PVDF) membranes (Millipore, Billerica, MA, USA). After blocking, the membranes were reacted with the appropriate primary antibodies. Immunostaining with secondary antibodies was completed and SuperSignal West Femto substrates (Pierce, Rockford, IL, USA) were used for detection.

Full paper   Login or join for free to view the full paper.

Reviews

Discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing Autophagy assay cell type - Human fetal osteoblastic (hFOB) 1.19 using Autophagy Inhibitor, 3-MA from Sigma-Aldrich.

Manufacturer protocol

Download the product protocol from Sigma-Aldrich for Autophagy Inhibitor, 3-MA below.

Download manufacturer protocol

Videos

Check out videos that might be relevant for performing Autophagy assay cell type - Human fetal osteoblastic (hFOB) 1.19 using Autophagy Inhibitor, 3-MA from Sigma-Aldrich. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.