Cell lysates (protein of 40 μg) were run on 10% SDS-polyacrylamide gel electrophoresis, subsequently transferred to nitrocellulose membrane. Blots were blocked with 5% BSA and then exposed to the appropriate primary antibody in dilutions (1:1000) suggested from the commercial supplier. Primary antibodies were probed with horseradish peroxidase conjugated goat anti-rabbit-IgG (SC-2004, Santa Cruz, CA, USA) or anti-mouse-IgG (SC-2005, Santa Cruz, CA, USA). Visualization was performed with enhanced chemiluminescence (RPN2209, Amersham, Buckinghamshire, UK). Capturing image was achieved by ChemiDoc image analyzer (Bio-Rad, Hercules, CA, USA).
Anti mouse igg sc 2005
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Anti-mouse IgG (sc-2005) is a secondary antibody product from Santa Cruz Biotechnology. It is designed to recognize and bind to mouse immunoglobulin G (IgG) antibodies.
Automatically generated - may contain errors
Lab products found in correlation
Anti rabbit igg sc 2004, by Santa Cruz Biotechnology (2 mentions)
Rpn2209, by Cytiva (1 mentions)
Chemidoc, by Bio-Rad (1 mentions)
Rabbit igg sc 2027, by Santa Cruz Biotechnology (1 mentions)
Rat anti ha clone 3f10, by Roche (1 mentions)
Mouse anti β actin clone ac 15, by Merck Group (1 mentions)
Mouse anti myc 9e10, by Santa Cruz Biotechnology (1 mentions)
Anti goat igg sc 2354, by Santa Cruz Biotechnology (1 mentions)
Rabbit anti p iκbα ser32, by Cell Signaling Technology (1 mentions)
Mouse anti v5, by Thermo Fisher Scientific (1 mentions)
Ab469, by Abcam (1 mentions)
Ab186733, by Abcam (1 mentions)
Vimentin 5741, by Cell Signaling Technology (1 mentions)
Ab32529, by Abcam (1 mentions)
Erlotinib, by Selleck Chemicals (1 mentions)
Anti gapdh sc 32233, by Santa Cruz Biotechnology (1 mentions)
Horseradish peroxidase conjugated anti rabbit sc 2004, by Santa Cruz Biotechnology (1 mentions)
Dulbecco s modified eagle s medium, by Corning (1 mentions)
Ly294002, by MedChemExpress (1 mentions)
Nci h460, by National Collection of Authenticated Cell Cultures (1 mentions)
6 protocols using anti mouse igg sc 2005
1
Western Blot Analysis Procedure
2
Antibody Characterization and Validation
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The following primary antibodies were used: rat anti-HA (clone 3F10, Roche), rabbit anti-MyD88 (#3699S), rabbit anti-p105 (#4717), rabbit anti-IκBα (44D4) (#4812S), rabbit anti-p-IκBα (Ser32) (#2859) (all from Cell Signaling), mouse anti-V5 (Thermo Scientific, #R96025), rabbit anti-BANK1 (HPA037002), mouse anti β-actin (clone AC-15) (A5441) (both from Sigma-Aldrich), mouse anti-BANK1 (F-8) (sc-393611), rabbit anti-TRAF6 (H-274) (sc-7221), mouse anti-TRAF6 (D-10) (sc-8409), goat anti-TLR7 (V-20) (sc-16245), goat anti-TLR9 (N-15) (sc-13215), and mouse anti-Myc (9E10) (sc-40) antibodies; as a nonspecific control, the immunoglobulins rabbit IgG (sc-2027) and mouse IgG (sc-2025) (all from Santa Cruz) were used. The HRP-linked secondary antibodies for western blotting were anti-rabbit IgG (sc-2004) (#7074), anti-mouse IgG (sc-2005) (#7076) (both from Santa Cruz and Cell Signaling), anti-goat IgG (sc-2354) or anti-rat IgG (sc-2032) (both from Santa Cruz). The secondary antibodies for IF reactions were Alexa Fluor 555 donkey anti-goat IgG (H + L) (#A-21432), Alexa Fluor 647 goat anti-mouse/rabbit IgG (H + L) (#A-21235; #A-31634) and Alexa Fluor 488 goat anti-rabbit/mouse IgG (H + L) (#A-11034; #A-11001) (all from Thermo Scientific).
3
Antibody Detection for EMT Markers
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Primary antibodies against AGO2 (ab186733) and Survivin (ab469) were purchased from Abcam Inc. (Cambridge, UK). Antibodies for detecting Snail (#3895) and Vimentin (#5741) were purchased from Cell Signaling Technology, Inc. (Danvers, MA, USA). Primary antibody for GAPDH (sc-25778) and secondary antibodies including anti-rabbit IgG (sc-2004) and anti-mouse IgG (sc-2005) were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA).
4
Lung Cancer Cell Line Culturing and Analysis
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The lung cancer cell lines NCI-H460 and NCI-H661 were purchased from the Type Culture Collection of the Chinese Academy of Sciences and were cultured in Dulbecco's modified Eagle's medium (Corning, Inc.) supplemented with 10% fetal bovine serum (Ausbian) in an incubator with 5% CO2 at 37˚C. Erlotinib was supplied by Selleck Chemicals, LY294002 was provided by MedChemExpress and monoclonal antibodies targeting p70S6K(ab32529) or anti-phospho-p70S6K (ab5231) were purchased from Abcam. Anti-GAPDH(sc-32233) and horseradish peroxidase-conjugated anti-rabbit (sc-2004)or anti-mouse IgG(sc-2005) were purchased from Santa Cruz Biotechnology, Inc.
5
Western Blot Analysis of Melanoma Signaling
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The tissue samples and total cell lysates were extracted with a 1X RIPA buffer and Western blot analysis was performed according to the methods of Lee et al.19 (link) Antibodies to phospho-ERK1/2 (p-ERK) (Cat. no. 9101), ERK (Cat. no. 9102), phospho-AKT (p-AKT) (Cat. no. 9271), AKT (Cat. no. 9272), caspase-3 (Cat. no. 14220), cleaved caspase-3 (Cat. no. 9664), PARP (Cat. no. 9542), cleaved PARP (Cat. no. 9541), Mcl-1 (Cat. no. 5453), Bcl-2 (Cat. no. 2870), Bax (Cat. no. 5023), and PUMA (Cat. no. 4976) were purchased form Cell Signaling Technology, Inc. Antibodies to Dock180 (SC-13163) and Rac1 (SC-217) were purchased from Santa Cruz Biotechnology, Inc. and antibodies to Elmo1 (ab2239) was purchased from Abcam. The secondary antibodies used were HRP conjugated anti-rabbit immunoglobulin (Ig) G (sc-2004), anti-goat IgG (sc-2020), and anti-mouse IgG (sc-2005) were purchased from Santa Cruz Biotechnology, Inc. The human malignant melanoma cells G361 were used as a positive control for antioxidant expression.
6
Global DNA Methylation Analysis in HEK293 Cells
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Genomic DNA was extracted from two replicates of DMSO or rotenone-treated HEK293 using 1:1:1 phenol: chloroform: isoamyl alcohol (Sigma, St. Louis, MO, United States). Global DNA methylation was first measured by dot blot analysis. Bisulfite-treated DNA (30–60 ng/μL) was denatured at 95°C for 5 min and then cooled at 4°C for 5 min in a conventional thermocycler (MyCycler; Bio-Rad; Hercules, CA, United States). DNA was spotted onto 0.45-micron nitrocellulose paper as 1- or 2-μL drops and dried for 30 min at room temperature. The membrane was UV cross-linked at 3000 Hz and incubated in anti-5methylcytosine (5mC) primary antibody overnight at 4°C (Epigentek 33D3; Farmingdale, NY, United States). The membrane was washed with TBST and incubated with a secondary antibody conjugated to HRP for 1 h at room temperature (Santa-Cruz Biotechnology anti-mouse IgG sc-2005; Dallas, TX, United States). The membrane was washed again with TBST after secondary incubation and visualized with chemiluminescence (ProSignal Femto; Prometheus; Raleigh, NC, United States). We used a serial dilution of 100% 5mC standard (Zymo Research D5012; Irvine, CA, United States) to create a standard ladder (Supplementary Figure 1 ).
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