KYSE-150 cells were treated with different dose of RP4010 for the indicated times. Cells were lysed with RIPA buffer (150 mM NaCl, 50 mM Tris-Cl, 1 mM EGTA, 1% Triton X-100, 0.1% SDS and 1% sodium deoxycholate, pH 8.0) supplemented with proteinase inhibitor cocktail (Sigma-Aldrich, US). Protein concentration was quantified using a BCA kit (Thermo, US). Primary antibodies used in this study included anti-STIM1 (1:500, BD Transduction), anti-Orai1 (1:1000, Millipore), anti-Cyclin B1 (1:1000, Cell Signaling Technology, US), anti-Cyclin D1 (1:1000, Cell Signaling Technology, US), anti-P27 (1:1000, Cell Signaling Technology, US), anti-STIM1 (1:1000, BD Transduction Laboratories, Clone 44), anti-Orai1 (1:1500, Millipore, against residues 22–40 of human protein, US), and anti-GAPDH (1:1000, GeneTex, US). Secondary antibodies included HRP-labeled goat anti-rabbit IgG (1:5000, Cell Signaling Technology, US) and anti-mouse IgG (1:5000, Cell Signaling Technology, US). Signals were detected on SpectraMax® i3 (Molecular Devices, CA).
Hrp labeled goat anti rabbit igg
Manufactured by Cell Signaling Technology
Sourced in United States
HRP-labeled goat anti-rabbit IgG is a secondary antibody conjugated with horseradish peroxidase (HRP). It is used to detect and visualize rabbit primary antibodies in various immunoassays.
Automatically generated - may contain errors
Lab products found in correlation
Alexa fluor 488 conjugated goat anti rabbit igg h l, by Thermo Fisher Scientific (5 mentions)
Bca kit, by Thermo Fisher Scientific (3 mentions)
Proteinase inhibitor cocktail, by Merck Group (3 mentions)
Hrp horseradish peroxidase labeled goat anti mouse igg, by Cell Signaling Technology (3 mentions)
β actin rabbit mab, by Cell Signaling Technology (3 mentions)
Plc γ1 pab, by Cell Signaling Technology (3 mentions)
Donkey anti goat igg h l hrp, by Abcam (3 mentions)
β tubulin rabbit pab, by ABclonal (3 mentions)
Horseradish peroxidase hrp conjugated goat anti mouse igg, by Cell Signaling Technology (2 mentions)
P plc γ1 ser1248 rabbit mab, by Cell Signaling Technology (2 mentions)
Egfr pab, by ABclonal (2 mentions)
Golga1 rabbit pab, by ABclonal (2 mentions)
Gp73 golph2 mouse mab, by Proteintech (2 mentions)
Alexa fluor 647 conjugated goat pab to rabbit igg, by Abcam (2 mentions)
Alexa fluor 488 conjugated donkey anti goat igg, by Abcam (2 mentions)
Alexa fluor 633 conjugated goat anti mouse igg h l, by Thermo Fisher Scientific (2 mentions)
Chemidoc, by Bio-Rad (2 mentions)
Anti egfr, by Thermo Fisher Scientific (2 mentions)
Anti β actin, by Proteintech (2 mentions)
Anti erbb2, by Thermo Fisher Scientific (2 mentions)
17 protocols using hrp labeled goat anti rabbit igg
1
Effects of RP4010 on Cell Cycle Regulators
2
Anti-inflammatory Mechanisms of Dexamethasone
Check if the same lab product or an alternative is used in the 5 most similar protocols
E. coli O111:B4 lipopolysaccharides (LPS) and dexamethasone were purchased from Sigma Aldrich (St. Louis, MO, USA). The primary antibodies used in this study, including anti-mouse iNOS (1:3,000 dilution), NF-kB p65 antibodies (1:200 dilution), and Alexa Fluor 488 anti-NF-kB p65 antibody, were obtained from Abcam (Cambridge, UK). The anti-β-actin antibody (1:1,000 dilution) was sourced from Cusabio (Wuhan, China). Horseradish peroxidase (HRP)-labeled goat anti-mouse IgG and HRP-labeled goat anti-rabbit IgG were purchased from Cell Signaling Technology (Beverly, MA, USA). Enzyme immunoassay reagents for the detection of TNF-α and IL-6 cytokines were obtained from BD Biosciences (San Jose, CA, USA).
+ Expand
E. coli O111:B4 lipopolysaccharides (LPS) and dexamethasone were purchased from Sigma Aldrich (St. Louis, MO, USA). The primary antibodies used in this study, including anti-mouse iNOS (1:3,000 dilution), NF-kB p65 antibodies (1:200 dilution), and Alexa Fluor 488 anti-NF-kB p65 antibody, were obtained from Abcam (Cambridge, UK). The anti-β-actin antibody (1:1,000 dilution) was sourced from Cusabio (Wuhan, China). Horseradish peroxidase (HRP)-labeled goat anti-mouse IgG and HRP-labeled goat anti-rabbit IgG were purchased from Cell Signaling Technology (Beverly, MA, USA). Enzyme immunoassay reagents for the detection of TNF-α and IL-6 cytokines were obtained from BD Biosciences (San Jose, CA, USA).
3
Immunoblot Analysis of Viral and Cellular Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used in this study: p-PLC-γ1(Ser1248) rabbit mAb (cat# 8713), PLC-γ1 pAb (cat# 2822S), β-actin rabbit mAb (cat# 4970), horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG (cat# 7076), as well as HRP-labeled goat anti-rabbit IgG (cat# 7074) were purchased from Cell Signaling Technology (Danvers, MA, USA). BoHV-1 gD mAb (cat# 1B8-F11) and goat anti-BoHV-1 serum (cat# PAB-IBR) were provided by VMRD Inc. (Pullman, WA, USA). EGFR pAb (cat# A11577) and GOLGA1 rabbit pAb (cat# A14688) were bought from Abclonal Technology (Woburn, MA, USA). GP73/GOLPH2 mouse mAb (cat# 66331-1-lg) was provided by Proteintech (Rosemont, IL, USA). Donkey anti-goat IgG H&L (HRP) (ca# ab97110), Alexa Fluor 647-conjugated goat pAb to rabbit IgG (cat# ab150079), and Alexa Fluor 488-conjugated donkey anti-goat IgG (cat# ab150129) were provided by Abcam (Cambridge, UK). Alexa Fluor 488-conjugated goat anti-rabbit IgG (H + L) (cat# A-11008) and Alexa Fluor 633-conjugated goat anti-mouse IgG (H + L) (Invitrogen, cat# A-21052) were provided by Invitrogen Life Technologies (Waltham, MA, USA).
4
EGFR/ERBB Expression in Esophageal Cell Lines
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Het-1A, KYSE-30, KYSE-150, KYSE-70 and KYSE-790 cell lines were cultured in a 6-well plate and harvest for Western blot. Cells were lysed with RIPA buffer (150 mM NaCl, 50 mM Tris-Cl, 1 mM EGTA, 1% Triton X-100, 0.1% SDS and 1% sodium deoxycholate, pH 8.0) and then supplemented with proteinase inhibitor cocktail (Sigma-Aldrich, Burlington, MA, USA). After that, protein concentrations were quantified by using a BCA kit (Thermo, Waltham, MA, USA). Primary antibodies used in this study included anti-EGFR (1:1000, Thermo, USA), anti-ERBB2 (1:1000, Thermo, Waltham, MA, USA), anti-ERBB3 (1:1000), anti-ERBB4 (1:1000) and anti-β-actin (1:1000, Proteintech, Rosemont, IL, USA). Secondary antibodies included HRP-labeled goat anti-rabbit IgG (1:5000) and anti-mouse IgG (1:5000, Cell Signaling Technology, Danvers, MA, USA). ECL substrate reagent (Cytiva Amersham, Marlborough, MA, USA) was used to visualize signals on ChemiDoc (BioRad, Hercules, CA, USA).
5
Immunoblot Analysis of Viral and Cellular Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used in this study: p-PLC-γ1(Ser1248) rabbit mAb (cat# 8713), PLC-γ1 pAb (cat# 2822S), β-actin rabbit mAb (cat# 4970), horseradish peroxidase (HRP)-conjugated goat anti-mouse IgG (cat# 7076), as well as HRP-labeled goat anti-rabbit IgG (cat# 7074) were purchased from Cell Signaling Technology (Danvers, MA, USA). BoHV-1 gD mAb (cat# 1B8-F11) and goat anti-BoHV-1 serum (cat# PAB-IBR) were provided by VMRD Inc. (Pullman, WA, USA). EGFR pAb (cat# A11577) and GOLGA1 rabbit pAb (cat# A14688) were bought from Abclonal Technology (Woburn, MA, USA). GP73/GOLPH2 mouse mAb (cat# 66331-1-lg) was provided by Proteintech (Rosemont, IL, USA). Donkey anti-goat IgG H&L (HRP) (ca# ab97110), Alexa Fluor 647-conjugated goat pAb to rabbit IgG (cat# ab150079), and Alexa Fluor 488-conjugated donkey anti-goat IgG (cat# ab150129) were provided by Abcam (Cambridge, UK). Alexa Fluor 488-conjugated goat anti-rabbit IgG (H + L) (cat# A-11008) and Alexa Fluor 633-conjugated goat anti-mouse IgG (H + L) (Invitrogen, cat# A-21052) were provided by Invitrogen Life Technologies (Waltham, MA, USA).
6
Quantitative Analysis of Apoptosis-Related Proteins
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The total protein content of cells or brain tissues was obtained using RIPA lysis buffer with protease inhibitor cocktail (Beyotime Institute of Biotechnology). The protein levels were quantified using a bicinchoninic acid protein assay kit (Beyotime Institute of Biotechnology) according to the manufacturer's instructions. The protein (20 µg/lane) was separated on 10% SDS-PAGE and transferred to a nitrocellulose membrane. The membranes were blocked using 5% skimmed milk at room temperature for 2 h and incubated overnight at 4˚C with the primary antibodies as follows: APC (1:2,000; cat. no. ab40778), NLRP3 (1:1,000; cat. no. ab263899), caspase-1 (1:100; cat. no. ab74279) (all from Abcam), gasdermin D (GSDMD-N; 1:1,000; cat. no. 39754) and β-actin (1:1,000; cat. no. 3700) (both from Cell Signaling Technology, Inc.)
After washing with 0.1 M PBS, the membranes were incubated with the secondary antibodies HRP-labeled goat anti-rabbit IgG (1:1,000, cat. no. A16104; Thermo Fisher Scientific, Inc.) and HRP-labeled goat anti-mouse antibody IgG (1:1,000, cat. no. A0216; Thermo Fisher Scientific, Inc) at 4˚C for 2 h. Enhanced chemiluminescence detection kit (cat. no. WBKLS0100; MilliporeSigma) was used for signal detection. Each reaction was replicated three times.
After washing with 0.1 M PBS, the membranes were incubated with the secondary antibodies HRP-labeled goat anti-rabbit IgG (1:1,000, cat. no. A16104; Thermo Fisher Scientific, Inc.) and HRP-labeled goat anti-mouse antibody IgG (1:1,000, cat. no. A0216; Thermo Fisher Scientific, Inc) at 4˚C for 2 h. Enhanced chemiluminescence detection kit (cat. no. WBKLS0100; MilliporeSigma) was used for signal detection. Each reaction was replicated three times.
7
MRGPRX2-mediated Activation Assay
8
Antibody Characterization for Cell Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The antibodies used in this study were as follows: 53BP1 polyclonal antibody (pAb) (Abcam, cat#ab87097), phosporylated-H2A.X (γH2A.X) monoclonal antibody (mAb) (Cell Signaling Technology, cat#2577), β-catenin mAb (Abcam, cat#ab32572), BoHV-1 gC mAb (VMRD, Inc., cat#F2), BoHV-1 gD mAb (VMRD, Inc., cat#1B8-F11), β-Actin mAb (ProteinTech Group, cat#60008-1-Ig), HRP (horseradish peroxidase)-labeled goat anti-mouse IgG (Cell Signaling Technology, cat#7076, 1:3000), HRP-labeled goat anti-rabbit IgG (Cell Signaling Technology, cat#7074), and Alexa Fluor 488®-conjugated goat anti-rabbit IgG (H + L) (Invitrogen, cat# A-11008).
9
Western Blot Antibody Validation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used in this study: CPT1A rabbit polyclonal antibody (pAb)(cat# A5307), β-Actin rabbit mAb (cat# AC026), and β-Tubulin Rabbit pAb(cat# AC015) were bought from Abclonal Technology (Woburn, MA, USA). COXIV rabbit pAb(cat#4844 S), HRP conjugated goat anti-mouse IgG (cat# 707 6), and HRP labeled goat anti-rabbit IgG (cat# 7074) were purchased from Cell Signaling Technology (Danvers, MA, USA). LaminA/C mouse mAb (cat# sc-376248) was provided by Santa Cruz Biotechnology (Dallas, TX, USA). BoHV-1 gD mAb (cat#1B8-F11), BoHV-1 gC mAb (cat#F2), and goat anti-BoHV-1 serum (cat# PAB-IBR) were provided by VMRD Inc. (Pullman, WA, USA). Donkey anti-goat IgG H&L (HRP) (ca#ab97110) was supplied by Abcam (Cambridge, UK). Alexa Fluor 488®-conjugated goat anti-rabbit IgG (H + L) (cat# A-11008) was provided by Invitrogen Life Technologies (Waltham, MA, USA). CPT1A-specific inhibitor Etomoxir (cat#HY-50202) was ordered from MedChemExpress (Monmouth Junction, NJ, USA).
10
Immunohistochemical Analysis of Transferrin Receptor
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To block endogenous peroxidase activity, we cut tumor tissues into 4 mm thick slices and incubated them with 3% hydrogen peroxide for 25 min at room temperature. Thereafter, the tissues were incubated with bovine serum albumin (BSA) for 30 min at room temperature to block nonspecific binding. The sections were incubated with primary antibodies (TFR, ab84036, 1:500; Abcam, Cambridge, UK) overnight at 4 °C and then incubated with a biotinylated secondary antibody (HRP-labeled goat anti-rabbit IgG, 1:200, Cell Signaling Technology, MA, USA) for 50 min at room temperature. The immunoreactions were visualized using a Dolichos Biflorus Agglutinin chromogenic reagent kit, which was followed by hematoxylin staining for 3 min. The sections were imaged using a fluorescence microscope (Leica DMI8) and analyzed with ImageJ (National Institutes of Health, Bethesda, MD, USA).
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!