Western blotting was performed as previously described.[43 ] Primary antibodies were VEGF‐A (EP1176Y, Abcam, USA) and Tubulin (11224‐1AP, Proteintech, USA) (loading control). Secondary antibody was anti‐rabbit IgG (7074s, CST, USA).
Tubulin 11224 1 ap
Manufactured by Proteintech
Sourced in United States, United Kingdom, China
Tubulin (11224-1-AP) is a primary antibody that specifically recognizes the tubulin protein. Tubulin is a key structural component of microtubules, which are essential for various cellular processes such as cell division, intracellular transport, and cell motility. This antibody can be used for applications such as western blotting, immunohistochemistry, and immunofluorescence to detect and analyze tubulin in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Polyvinylidene difluoride membrane, by Merck Group (2 mentions)
Sds polyacrylamide gel electrophoresis sds page, by Bio-Rad (2 mentions)
Phosphatase inhibitor cocktail, by Merck Group (2 mentions)
Bca assay, by Thermo Fisher Scientific (2 mentions)
Anti raga d8b5, by Cell Signaling Technology (2 mentions)
Anti aid maid 2, by Thermo Fisher Scientific (2 mentions)
Lamin b 66095 1 ig, by Proteintech (2 mentions)
Tfe3 hpa023881, by Merck Group (2 mentions)
Anti tfeb a303 673a, by Fortis Life Sciences (2 mentions)
Anti raptor 24c12, by Cell Signaling Technology (2 mentions)
Enhanced chemi luminescence (ecl), by Thermo Fisher Scientific (2 mentions)
Protease inhibitor cocktail, by Merck Group (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Normal igg peroxidase conjugated affinipure goat anti rabbit mouse igg h l, by Jackson ImmunoResearch (1 mentions)
β actin 60008 1 lg, by Proteintech (1 mentions)
Gsk3β 22104 1 ap, by Proteintech (1 mentions)
Gst 10000 0 ap, by Proteintech (1 mentions)
Ha no 3724, by Cell Signaling Technology (1 mentions)
Mg132, by Selleck Chemicals (1 mentions)
E cadherin 3195, by Cell Signaling Technology (1 mentions)
8 protocols using tubulin 11224 1 ap
1
Western Blotting of VEGF-A and Tubulin
2
Immunoblotting Assay with Proteasome and Protein Synthesis Inhibitors
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The proteasome inhibitor MG132 and protein synthesis inhibitor cycloheximide (CHX) were purchased from Selleck. Antibodies used in immunoblotting: eEF2K (ab45168, ab85721, 1:1000) was purchased from Abcam; PD-L1 (17952-1-AP, 1:1000), GSK3β (22104-1-AP, 1:2000), β-actin (60008-1-lg, 1:5000), Flag (66008-3-lg, 1:5000), tubulin (11224-1-AP, 1:5000), GST (10000-0-AP, 1:4000) were purchased from Proteintech; HA (No. 3724, 1:1000), p-GSK3β/S9 (No. 9323, 1:1000) were purchased from Cell Signaling Technologies; phosphoserine monoclonal antibody (mAb) (CSB-MA080235, 1:1000) was purchased from CUSABIO. Normal IgG/Peroxidase-conjugated AffiniPure Goat Anti-Rabbit/Mouse IgG (H+L) was purchased from Jackson Immuno Research.
3
Colorectal Cancer Cell Culture and Protein Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The SW480 and HT-29 cells were purchased from the cell bank of Shanghai Institutes for Biological Sciences (Shanghai, China). SW480 were cultured in RPMI 1640 (Hyclone, Logan, TX, USA) and HT-29 were cultured in DMEM (Gibco, Brookyln, NY, USA) with 5% CO2 in 37 °C. All media were supplemented with 10% Fetal Bovine Serum (16000-044, Gibco, Brookyln, NY, USA) and 1% penicillin/streptomycin. The main antibodies including purchased from PHF19 (77271), Cyclin D1 (55506), E-cadherin (3195), N-cadherin (13116), β-catenin (8480), and Twist (46702) were purchased from Cell Signaling Technology (Beverly, MA, USA); H3K27me3 (ab6002) was purchased from Abcam (Cambridge, UK); CDK4 (11026-1-AP), CDK6 (14052-1-AP), Tubulin (11224-1-AP), and GADPH (10494-1-AP) were purchased from Proteintech (Wuhan, Hubei, China).
4
Cell Fractionation and Immunoblotting Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Whole-cell lysates and separate nuclear/cytoplasmic fractions were extracted from cells according to routine protocols. Western blotting and coimmunoprecipitation were executed as preceding description [21 (link)]. The antibodies against GAPDH (60004-1-Ig), Flag-tag (20543-1-AP), α-tubulin (11224-1-AP), Lamin B1 (12987-1-AP), and GST-tag (HRP-66001) were purchased from ProteinTech. The antibodies against TNPO1 (ab10303, Abcam) and FUBP1 (ab181111, Abcam).
5
Immunoprecipitation and Immunoblotting Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
PPARγ (16643‐1‐AP), Tubulin (11224‐1‐AP), PD‐L1 (66248–1), ATG7 (10088‐2‐AP), LC3B (14600‐1‐AP), GST (66001‐2), Lamp1(21997‐1‐AP), and Flag (66008‐4) antibodies were obtained from Proteintech. Secondary antibodies were obtained from Jackson Immunoresearch. Cells were lysed in lysis buffer (50 mM Tris–HCl pH 7.4, 250 mM NaCl, 0.5% Triton X100, 10% glycerol, 1 mM DTT, protease inhibitor cocktail). Protein concentration in the supernatant was determined by the Pierce BCA Protein Assay Kit (Thermo). For immunoprecipitation, cell lysates were precleared with protein A/G magnetic bead (Cat: B23202; Bimake). Precleared lysates were subjected to immunoprecipitation using the indicated antibodies with protein A/G magnetic beads as described previously.49 , 50 The samples were subjected to SDS‐PAGE and immunoblotted with indicated antibodies. Data are triplicates from three independent experiments. Blots were quantified by Image J.
6
Western Blot Analysis of Signaling Pathways
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
B cells were lysed in radioimmunoprecipitation assay (RIPA) buffer (50 mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate (SDS)) supplemented with protease inhibitor cocktail and phosphatase inhibitor cocktail (Sigma-Aldrich). Protein concentration was detected by BCA assay (Thermo Fisher Scientific), and an equal amount of protein was resolved in 4–12% SDS-polyacrylamide gel electrophoresis (SDS-PAGE) (Bio-Rad). Proteins were transferred to polyvinylidene difluoride membranes (Millipore) and probed overnight with the following primary antibodies: anti-p-S6K (108D2), anti-p-S6 (D57.2.2E), anti-LAMP1(C54H11), anti-p-4EBP1 (236B4), anti-Raptor (24C12), and anti-RagA (D8B5) all from Cell Signaling Technology), anti-AID (mAID-2, Thermo Fisher Scientific), anti-TFEB (A303–673A, Bethyl Laboratories), Lamin B (66095–1-Ig, Proteintech), tubulin (11224–1-AP, Proteintech), TFE3 (HPA023881, Sigma-Aldrich) and anti-b-actin (13E5, Sigma-Aldrich). The membrane was washed and incubated with indicated secondary antibody for the subsequently enhanced chemiluminescence (ECL, Thermo Fisher) exposure.
7
Western Blot Analysis of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Radio immunoprecipitation assay buffer (R0010, Solarbio, Beijing, China) was used to lyse the cells for 20 min to extract total protein. The protein concentrations were detected using the Bio-Rad Protein Assay Kit (PC0020, Servicebio, Wuhan, China). Then the proteins were isolated with 12% SDS-PAGE and retransferred onto PVDF membranes (Meilunbio, Dalian, China). The membranes were blocked with 5% skimmed milk for 2 h and incubated with primary antibodies, including cyclin D1 (26939-1-AP), cyclin E (11554-1-AP), C-myc (10828-1-AP), CDK4 (11026-1-AP), p38 (14064-1-AP), p-P38 (Thr180/Tyr182)-(28796-1-AP), ERK (16443-1-AP), p-ERK (Thr202/Tyr204)-(28733-1-AP), AKT (10176-2-AP), p-AKT (Ser473)-(66444-1-Ig), Bcl-2 (60178-1-lg), and tubulin (11224-1-AP), which were purchased from Proteintech Company (Wuhan, China). JNK (9252) and p-JNK (Thr183/Tyr185)-(9255) were obtained from Cell Signaling Technology (Danvers, MA, USA). Cleaved-caspase3 (A11021) was obtained from ABclonal (Wuhan, China). After washing three times with tris-buffered saline, the secondary antibodies were incubated for 2 h. Band intensity was measured using a chemiluminescence Ultra High Sensitivity ECL Kit (AR1190, Boster, Wuhan, China) and analyzed by the ImageJ software (ImageJ 6.0, Bethesda, MD, USA).
8
Quantitative Western Blot Analysis of B Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
B cells were lysed in radioimmunoprecipitation assay (RIPA) buffer (50 mM Tris (pH 7.4), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate, 0.1% sodium dodecyl sulfate (SDS)) supplemented with protease inhibitor cocktail and phosphatase inhibitor cocktail (Sigma-Aldrich). Protein concentration was detected by BCA assay (Thermo Fisher Scientific), and an equal amount of protein was resolved in 4–12% SDS-polyacrylamide gel electrophoresis (SDS-PAGE) (Bio-Rad). Proteins were transferred to polyvinylidene difluoride membranes (Millipore) and probed overnight with the following primary antibodies: : anti-p-S6K (108D2), anti-p-S6 (D57.2.2E), anti-LAMP1(C54H11), anti-p-4EBP1 (236B4), anti-Raptor (24C12), and anti-RagA (D8B5) all from Cell Signaling Technology), anti-AID (mAID-2, Thermo Fisher Scientific), anti-TFEB (A303–673A, Bethyl Laboratories), Lamin B (66095–1-Ig, Proteintech), tubulin (11224–1-AP, Proteintech), TFE3 (HPA023881, Sigma-Aldrich) and anti-β-actin (13E5, Sigma-Aldrich). The membrane was washed and incubated with indicated secondary antibody for the subsequently enhanced chemiluminescence (ECL, Thermo Fisher) exposure.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!