The Protease Inhibitor Cocktail Set Ⅲ DMSO Solution (EDTA Free), 4-(2-Aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF), aprotinin bovine lung, and leupeptin hemisulfate monohydrate were obtained from Wako. The stocks were dissolved in water. Bestatin and pepstatin A were also obtained from FUJIFILM Wako Pure Chemical Corporation, and the stocks were dissolved in DMSO. E-64 was obtained from Sigma-Aldrich, and the stock was dissolved in water. Each protease was used with the following final concentration : AEBSF 1 mmol / L, aprotinin bovine lung 0.8 µmol / L, E-64 15 µmol / L, leupeptin hemisulfate 20 µmol / L, Bestatin 50 µmol / L, pepstatin A 10 µmol / L.
Bestatin
Manufactured by Fujifilm
Bestatin is a laboratory equipment product manufactured by Fujifilm. It is a potent and selective inhibitor of aminopeptidase B, an enzyme involved in the metabolism of various peptides and proteins.
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Lab products found in correlation
2 protocols using bestatin
1
Protease Inhibitor Cocktail for Experiments
2
Western Blotting of B16F10 Cells
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B16F10 cells were seeded in a 6-cm dish at 4 × 105 cells/dish and cultured with α-mg. The cells were washed twice with RIPA buffer (50 mM Tris-HCl [pH 7.4], 150 mM NaCl, 1% Triton X-100, 0.1% SDS, 0.5% Na-deoxycholate, 1 mM EDTA, 10 mM NaF, 1 mM Na3VO4, 0.8 μM Aprotinin [Wako], 50 μM Bestatin [Wako], 20 μM Leupeptin [Wako], 10 μM Pepstatin A [Wako] and 1 mM AEBSF [Wako]). Then, cells were disrupted using an ultrasonic homogenizer and centrifuged to obtain a supernatant for western blotting. After the antibody reaction, the membrane was treated with LuminoGLO reagent (Cell Signaling Technology) to cause the band of the target protein to emit light, and the band was detected and photographed using AE-9300H Ez-Capture MG (ATTO Corporation, Tokyo, Japan). Each protein detection was performed at least 3 times replication.
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