Cells were grown to 90% confluence on 6‐cm dishes, washed with PBS, lysed and protein was recovered by sonication with RIPA buffer (Thermo Scientific). Dynabeads‐protein G for immunoprecipitation (Life Technologies) were incubated with the primary antibodies (anti‐EP4 [Cayman] and anti‐Orai1 [Sigma‐Aldrich]) or (anti‐EP4 [Cayman] and anti‐TRPC1 [Abcam]) for 24 hours at 4°C.21 , 26 These antibody‐coated Dynabeads (Life Technologies) bound to the target proteins were separated by a magnet, and after repeated washing four times, the isolated protein complexes were subjected to western blotting with the respective antibodies.
Anti trpc1
Manufactured by Abcam
Sourced in United States
Anti-TRPC1 is a primary antibody that specifically binds to the TRPC1 protein. TRPC1 is a member of the transient receptor potential (TRP) cation channel subfamily C. The core function of this product is to detect and quantify TRPC1 expression in various biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Anti cdk2, by Cell Signaling Technology (2 mentions)
Anti pi3k p85α, by Bioworld Technology (2 mentions)
Anti calmodulin, by Santa Cruz Biotechnology (2 mentions)
Anti gapdh, by Cell Signaling Technology (2 mentions)
Anti cyclin a, by Santa Cruz Biotechnology (2 mentions)
Anti cdk6, by Cell Signaling Technology (2 mentions)
Anti orai1, by Merck Group (1 mentions)
Anti ep4, by Cayman Chemical (1 mentions)
Dynabeads protein g for immunoprecipitation, by Thermo Fisher Scientific (1 mentions)
Ripa buffer, by Thermo Fisher Scientific (1 mentions)
Dynabead, by Thermo Fisher Scientific (1 mentions)
Anti akt, by Cell Signaling Technology (1 mentions)
Anti cdk1, by Cell Signaling Technology (1 mentions)
Anti erk1 2, by Cell Signaling Technology (1 mentions)
Anti p erk1 2 thr202 tyr204, by Cell Signaling Technology (1 mentions)
Anti p21cip1, by Cell Signaling Technology (1 mentions)
Anti p akt ser473, by Cell Signaling Technology (1 mentions)
Ax70 microscope, by Olympus (1 mentions)
Anti β actin, by Abcam (1 mentions)
Anti p erk, by Abcam (1 mentions)
5 protocols using anti trpc1
1
Immunoprecipitation of EP4 and Orai1/TRPC1
2
Western Blot Analysis of Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins were extracted, determined, and separated by the SDS-page technique as previously described [44 (link)]. The primary antibodies used were: anti-TRPC1 (1:1000, Abcam, Waltham, MA, USA), anti-GAPDH (1:2000, Cell Signaling Tech., Danvers, MA, USA), anti-CDK6 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-Cyclin A (1:500, Santacruz Biotechnology, Dallas, TX, USA), anti-CDK2 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-CDK1 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-p21CIP1 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-PI3K p85α (1:500, Bioworld Technology, tebu-bio, France), anti-calmodulin (1:500, Santacruz Biotechnology, Dallas, TX, USA), anti-pAKT (Ser473) (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-AKT (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-pERK1/2 (Thr202/Tyr204) (1:500, Cell Signaling Tech., Danvers, MA, USA), and anti-ERK1/2 (1:500, Cell Signaling Tech., Danvers, MA, USA). Secondary antibodies (1:4000, Cell Signaling Tech., Danvers, MA, USA) were coupled to horseradish peroxidase, and proteins were detected using enhanced chemiluminescence (Ozyme, Saint-Cyr-l’Ecole, France). Quantification was performed with the ImageJ software 1.53a (National Institute of Health, Bethesda, MD, USA) analysis tool. All experiments were normalized to the level of GAPDH.
3
Renal Fibrosis Biomarker Evaluation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Kidneys were fixed with 10% neutral buffered formalin at room temperature for 3 h and processed for immunostaining using standard techniques. Expression levels of collagen type 1 (Col1), GRP78, TRPC1 and PERK were measured by immunohistochemical staining methods. Paraffin-embedded sections (3-mm thick) were mounted on poly-L-lysine-coated glass slides overnight at 4°C. Subsequently, sections were incubated at 37°C for 1 h with the following primary antibodies: Anti-Col1 (cat. no. ab34710; 1:1,000; Abcam), anti-GRP78 (cat. no. ab21685; 1:1,000; Abcam), anti-TRPC1 (cat. no. ab51255; 1:1,000; Abcam), anti-PERK (cat. no. ab65142; 1:1,000; Abcam) and anti-β-actin (cat. no. ab8227; 1:1,000; Abcam). Sections were incubated with horseradish peroxidase-conjugated di-antibody (cat. no. HAF008; 1:2,000; R&D Systems). Subsequently, chromogen detection was performed using 3,3-DAB. For the control test, the binding antibody was omitted and PBS was used instead. Images were captured using a VKC150 color video camera (Hitachi, Ltd.) and an AX70 microscope (Olympus Corporation).
4
Protein Expression Analysis in Pulmonary Arterial Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The total protein extracted from deendothelialized distal PA homogenate was subjected to 12% SDS-PAGE and then transferred to a polyvinylidene fluoride membrane (Millipore, MA). Following blockade with 5% BSA buffer for 2 h, membranes were probed with anti-STIM2 (1:200; Alomone Labs, Jerusalem, Israel), anti-Orai1 (1:200; Alomone Labs, Jerusalem, Israel), anti-Orai2 (1:200; Alomone Labs, Jerusalem, Israel), anti-TRPC1 (1:300; Abcam, MA), anti-TRPC4 (1:200; Alomone Labs, Jerusalem, Israel), anti-β-actin (1:500; Cell Signalling Technology, MA) primary antibodies overnight and horseradish peroxidase (HRP)-conjugated secondary antibodies (1:5000; Cell Signalling Technology, MA) for 1 h. The bands were visualized by enhanced chemiluminescence reagent (Thermo Fisher Scientific, CA). The optical density of each blot was quantified using Quantity One software (Bio-Rad, CA) and normalized to β-actin on the same membrane.
5
Immunoblotting for Cell Signaling Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Proteins were extracted and separated as previously described [60 (link)]. The primary antibodies used were: anti-TRPC1 (1:1000, Abcam, Waltham, MA, USA), anti-αTubulin (Sigma-Aldrich, France), anti-CDK6 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-Cyclin A (1:500, Santacruz Biotechnology, Dallas, TX, USA), anti-CDK2 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti-p21 (1:500, Cell Signaling Tech., Danvers, MA, USA), anti- GAPDH (1:2000, Cell Signaling Tech., Danvers, MA, USA), anti-PI3K p85α (1:500, Bioworld Technology), and anti-Calmodulin (1:500, Santacruz Biotechnology, Dallas, TX, USA). Secondary antibodies were coupled to horseradish peroxidase, permitting protein detection with an enhanced chemiluminescence kit (Ozyme, Saint-Cyr-l’Ecole, France). Quantification was performed with the ImageJ analysis tool. All experiments were normalized to α-Tubulin or GAPDH, which were used as reference proteins.
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!