Neuroblastoma tumor cell lines were obtained from the University of California San Francisco Cell Culture Facility (Kelly, SK-N-BE2, and SH-EP). SMS-KCN, SHEP MYCNwt and MYCNT58A/S62A cells were obtained from Martin Eilers lab. All cells were grown in RPMI with 10% FBS. Neuroblastoma cells were harvested and lysed with Cell Signaling Lysis buffer + 1% SDS, sonicated and supernatants boiled in LDS sample buffer (Invitrogen). Western blots were performed as described previously (Chanthery et al., 2012 ), with primary antibodies to MYCN (ab24193, Abcam), Histone H3, P-Histone H3 (S10), Aurora A (Cell signaling), and GAPDH (Millipore). Western blot quantitation performed with ImageJ software. VX-680 (S1048) and MLN8237 (S1133) were obtained from Selleck chemicals.
Ab24193
Manufactured by Abcam
Ab24193 is a lab equipment product offered by Abcam. It serves a core function, but a detailed description cannot be provided while maintaining an unbiased and factual approach. Therefore, the description for this product is not available.
Automatically generated - may contain errors
Lab products found in correlation
Lds sample buffer, by Thermo Fisher Scientific (1 mentions)
Gapdh, by Merck Group (1 mentions)
P histone h3 s10, by Cell Signaling Technology (1 mentions)
Vx 680 s1048, by Selleck Chemicals (1 mentions)
Aurora a, by Cell Signaling Technology (1 mentions)
Ripa lysis buffer, by Thermo Fisher Scientific (1 mentions)
Enhanced chemiluminescence, by Thermo Fisher Scientific (1 mentions)
Protease inhibitor cocktail, by Thermo Fisher Scientific (1 mentions)
Fusion solo, by Vilber (1 mentions)
2 protocols using ab24193
1
Neuroblastoma cell line characterization
2
Protein Extraction and Western Blot Analysis
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Proteins were extracted from harvested cells using RIPA lysis buffer (Thermo Fisher Scientific, #89,900) containing a protease inhibitor cocktail (Thermo Fisher Scientific, #78,430). The total protein concentration of each cell lysate was measured using the Bradford method, with bovine serum albumin as a standard. Equal amounts of protein lysate (45 µg) were loaded onto denaturing polyacrylamide gels and transferred to a nitrocellulose membrane. The primary antibodies used for immunoblotting were anti-RAB27A (diluted 1:2000; #ab55667; Abcam), anti-RAB27B (diluted 1:2000; Cat # PA5-54,096; Thermo Fisher Scientific), and anti-MYCN (diluted 1:500; Cat # ab24193; Abcam) followed by incubation with horseradish peroxidase-conjugated secondary antibodies. The immunoreactive bands were detected using enhanced chemiluminescence (Cat #32,109; Thermo Fisher Scientific). Digital chemiluminescent images were captured and analyzed using Fusion Solo (Vilber, Marne-la-Vallee Cedex, France).
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