Cell lysates were mixed with SDS buffer (3 M, Maplewood, Minnesota, USA), and denatured at 100 °C for 5 min using a standard protocol. Adequate amount of proteins (30 µg) were separated using 10% SDS-polyacrylamide gel electrophoresis and blotted onto nitrocellulose membranes (Whatman, Dassel, Germany). The membranes were blocked with 5% non-fat skim milk in TBS-T buffer, followed by incubation with primary antibodies in 5% skim milk. Primary antibodies, such as anti-MITF, anti-CREB, anti-tyrosinase, anti-TRP-2, anti-p-CREB, anti-TRP-1, and anti-β-actin were purchased from Bioworld Technology (St. Louis Park, MN, USA). Anti-mouse IgG-horseradish peroxidase (HRP) and anti-goat IgG-HRP from Santa Cruz Biotechnology was used as secondary antibody. An ECL solution system (Perkin Elmer) was used to detect the antigen-antibody reaction.
Anti mitf
Manufactured by Bioworld Technology
Sourced in United States
Anti-MITF is a laboratory reagent used to detect and quantify the MITF (Microphthalmia-associated transcription factor) protein in biological samples. MITF is a key regulator of melanocyte development and function, and its expression is often altered in various skin conditions and melanoma. Anti-MITF provides a tool for researchers to study MITF-related pathways and mechanisms.
Automatically generated - may contain errors
Lab products found in correlation
Nitrocellulose membrane, by Cytiva (3 mentions)
Anti tyrosinase, by Bioworld Technology (3 mentions)
Ecl solution system, by PerkinElmer (2 mentions)
Anti trp 2, by Bioworld Technology (2 mentions)
Anti trp 1, by Bioworld Technology (2 mentions)
Anti tyrp 2, by Bioworld Technology (2 mentions)
Anti goat igg horse radish peroxidase, by Santa Cruz Biotechnology (2 mentions)
Anti mouse igg hrp, by Santa Cruz Biotechnology (2 mentions)
Anti mouse igg horseradish peroxidase hrp, by Santa Cruz Biotechnology (1 mentions)
Anti goat igg hrp, by Santa Cruz Biotechnology (1 mentions)
Anti β actin, by Bioworld Technology (1 mentions)
Arbutin, by Merck Group (1 mentions)
Thiazolyl blue tetrazolium bromide, by Merck Group (1 mentions)
2 2 azobis 2 amidinopropane dihydrochloride, by Merck Group (1 mentions)
Hydroquinone, by Merck Group (1 mentions)
8 methoxypsoralen 8 mop, by Merck Group (1 mentions)
Anti phospho p38, by Bioworld Technology (1 mentions)
Anti p38, by Bioworld Technology (1 mentions)
O tetradecanoyl phorbol 13 acetate, by Merck Group (1 mentions)
L dopa, by Merck Group (1 mentions)
4 protocols using anti mitf
1
Western Blot Analysis of Melanogenic Proteins
2
Melanogenesis Regulation in Vitro
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Arbutin, 3-isobutyl-1-methylxanthine (IBMX), L-DOPA (L-3,4-dihydroxyphenylalanine), sodium hydroxide, thiazolyl blue tetrazolium bromide (MTT), O-tetradecanoyl phorbol-13-acetate (TPA), 2,2’-azobis(2-amidinopropane) dihydrochloride (AAPH), 8-methoxypsoralen (8-MOP), 3-Isobutyl-1-methyl-2,6(1H,3H)-purinedione (IBMX), and hydroquinone were obtained from Sigma-Aldrich Co. (St. Louis, MO, USA). All other reagents and chemicals were high-grade and commercially available. Antibodies bought from Bioworld Technology (St. Louis Park, MN, USA) included anti-Tyrosinase, anti-TYRP-1, anti-TYRP-2, anti-MITF, anti-phospho-p38, and anti-p38, anti-phospho-ERK, and anti-ERK, anti-phospho-JNK, and anti-JNK.
3
Melan-a Cell Protein Expression Analysis
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Melan-a cell lysates were mixed with sample buffer (250 mM Tris-HCl (pH 6.8), 0.5 M DTT, 10% SDS, 0.5% bromophenol blue, 50% glycerol, 5% 2-mercaptoethanol), and denatured at 100 °C for 5 min using a standard protocol. A 10% of SDS-PAGE was used to separate the sample proteins (50 µg). Following electrotransfer to nitrocellulose membranes (Whatman, Dassel, Germany), the membranes were immersed overnight in a mixture containing antibodies and 5% skim milk. Primary antibodies, such as anti-tyrosinase, anti-TRP-1, anti-TRP2, anti-MITF, p-CREB, and total CREB and β-actin, were from Bioworld Technology (St. Louis Park, MN, USA). Secondary antibodies (anti-goat IgG-horse radish peroxidase (HRP) and anti-mouse IgG-HRP) were purchased from Santa Cruz. The resulting reaction was exposed using an ECL solution system (Perkin Elmer).
4
Melanoma Cell Protein Expression
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Melan-a cell lysates were prepared using a standard protocol, mixed with 5X SDS-PAGE (3M Science, Seoul, Korea) sample buffer and denatured at 100 °C for 5 min. An equal amount (20 µg protein) of each sample was separated by 10% SDS-PAGE gel electrophoresis, followed by electrotransfer to nitrocellulose membranes (Whatman, Dassel, Germany). The membranes were then incubated overnight with 5% skim milk (for anti-Tyr, anti-TYRP-1, anti-TYRP-2, anti-MITF) or 5% bovine serum albumin (BSA) for anti-phospho-JNK, anti-JNK, anti-phospho-p38, anti-p38, and Anti-p44/42MAPK (ERK1/2), and anti-phospho-p44/42MAPK (ERK1/2), detection. anti-Tyr, anti-TYRP-1, anti-TYRP-2, anti-MITF (Bioworld Technology, St. Louis Park, MN, USA) for Western blotting, and anti-phospho-JNK, anti-JNK, anti-phospho-p38, anti-p38, and Anti-p44/42MAPK (ERK1/2), and anti-phospho-p44/42MAPK (ERK1/2), and anti-MITF (Cell Signaling Technology, Beverly, MA, USA) for signaling studies were utilized as 1st antibodies. Anti-goat IgG-horse radish peroxidase (HRP) and anti-mouse IgG-HRP were purchased from Santa Cruz and used as 2nd antibodies. The reaction was proceeded using an ECL system (Perkin Elmer).
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