Protein lysates were prepared in LDS sample buffer and separated using the Bolt SDS-PAGE system on 12% Bis-Tris gels, before transfer to nitrocellulose membranes (all Life Technologies). Protein expression was analyzed using the following antibodies: mouse monoclonal acetyl-Lysine, mouse α-Tubulin, rabbit GAPDH, rabbit Glutamate Dehydrogenase (GDH), rabbit acetyl-Tubulin K40, rabbit HDAC6, rabbit LC3A/B, rabbit COX IV, rabbit Succinate Dehydrogenase A (SDHA), rabbit phospho-ACC from Cell Signaling Technologies; rabbit PINK1 from Novus Biochemicals. Fluorescent anti-mouse or anti-rabbit secondary antibodies (red, 700 nm; green, 800 nm) from LiCor were used to detect expression levels.
Mouse α tubulin
Manufactured by Cell Signaling Technology
Sourced in United States
Mouse α-Tubulin is a primary antibody that recognizes the α-tubulin protein, a key component of the cytoskeleton in mammalian cells. It is used for the detection and localization of α-tubulin in various applications, such as immunofluorescence, western blotting, and immunohistochemistry.
Automatically generated - may contain errors
Lab products found in correlation
Ripa lysis buffer, by Thermo Fisher Scientific (2 mentions)
Nitrocellulose membrane, by Thermo Fisher Scientific (1 mentions)
Mouse monoclonal acetyl lysine, by Cell Signaling Technology (1 mentions)
Rabbit lc3a b, by Cell Signaling Technology (1 mentions)
Fluorescent anti mouse or anti rabbit secondary antibodies, by LI COR (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Thermo Fisher Scientific (1 mentions)
Halt protease and phosphatase inhibitor cocktail, by Thermo Fisher Scientific (1 mentions)
Odyssey infrared imaging system, by LI COR (1 mentions)
Irdye 680, by LI COR (1 mentions)
Irdye 800, by LI COR (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Imagequant las 4000 mini biomolecular imager, by Cytiva (1 mentions)
Rabbit parp 1, by Abcam (1 mentions)
Pvdf membrane, by Bio-Rad (1 mentions)
Chemi lumi one super, by Nacalai Tesque (1 mentions)
Rabbit phospho erk 1 2, by Cell Signaling Technology (1 mentions)
Las 4000 mini, by GE Healthcare (1 mentions)
M per mammalian protein extraction reagent, by Thermo Fisher Scientific (1 mentions)
4 protocols using mouse α tubulin
1
Western Blot Analysis of Cellular Proteins
2
Quantifying COVID-19 Platelet Signaling
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Protein levels of pAKT, AKT and PI3K in platelets from COVID‐19 patients and healthy controls were determined by western blot (WB). In brief, following platelet isolation, cells were centrifuged (5 min, 700 g at 4°C) and the platelet pellet resuspended in ice‐cold RIPA lysis buffer (ThermoFisher Scientific) containing Halt™ protease and phosphatase inhibitor cocktail (100×; ThermoFisher Scientific) and EDTA (0.5 M, 100× ThermoFisher Scientific). The proteins were separated by electrophoresis using 12% SDS‐PAGE gels in glycine‐tris buffer and subsequently transferred to polyvinylidene difluoride (PVDF) membranes (0.45 µm; Merck). Finally, the membranes were incubated with rabbit phospho‐AKT antibody (1:1000; Invitrogen), mouse AKT antibody (1:1000; Abcam, Signaling), rabbit pPI3K (p85,N‐SH, 1:1000; Merck) and mouse α‐Tubulin (Cell Signaling) which was followed by the incubation with appropriate secondary anti‐rabbit IRDye®800 and anti‐mouse antibody IRDye®680 (1:3000, LI.COR®). Protein bands were detected with Odyssey infrared imaging system (LI.COR®). Relative intensity of western blots was quantified using ImageJ software (NIH).
3
Quantitative Western Blotting Analysis
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Western blotting was performed as previously described (21 (link)). Briefly, cells were lysed in Laemmli's buffer. Total proteins were separated using SDS-PAGE and were then transferred to PVDF membranes (Bio-Rad Laboratories, Inc.). After blocking, the membranes were incubated with primary antibodies against rabbit phosphorylated (p)CREB (Ser133; 1:1,000, cat. no. ab32096; Abcam), rabbit PARP-1 (1:1,000 cat. no. 9542; Cell Signaling Technology, Inc.) and mouse α-tubulin (1:1,000, cat. no. 3873; Cell Signaling Technology, Inc.) which was followed by incubation with appropriate horseradish peroxidase-conjugated secondary antibodies against rabbit (1:2,000, cat. no. p0448; Dako Agilent Technologies) and mouse (1:2,000, cat. no. p0260; Dako Agilent Technologies). The expression was visualized using an ECL detection kit (cat. no. RPN2106; Cytiva). Images were acquired using ImageQuant LAS 4000 Mini biomolecular imager (Cytiva). Semi-quantification on the relative expression of proteins was performed using ImageJ 2.1.0 software (National Institutes of Health).
4
Western Blot Analysis of Cellular Proteins
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The cells were washed twice with PBS, and lysed using M-PER mammalian protein extraction reagent (Thermo Fisher Scientific, USA) with protease (Thermo Fisher Scientific, USA) and phosphatase (Nacalai Tesque, Japan) inhibitors. For western blot analysis, proteins were separated by SDS-PAGE and transferred onto PolyVinylidene DiFluoride (PVDF) membranes. The membranes were blocked with 5% non-fat dry milk and incubated overnight at 4 °C using the following primary antibodies: mouse V5 (Nacalai Tesque, USA), rat FLAG (Novus Biologicals, USA), rabbit ERK1/2, rabbit phospho-ERK1/2, mouse α-Tubulin (Cell Signaling Technology, USA), or mouse α smooth muscle actin (SMA) (Sigma-Aldrich, USA). Bands were detected using Chemi-Lumi One Super (Nacalai Tesque, Japan) and Chemiluminescence detection system LAS-4000 Mini (GE Healthcare, USA).
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