Vero cells transfected with CRISPR/Cas9 plasmid targeted ICP0 gene, infected with HSV-1 at 0.1 PFU/cell after 3 days. At 4 h and 24 h after infection, cells were fixed with methanol for 20 min at −20°C, washed with PBS, layered with mAbs to the ICP0 protein of HSV-1 (Anti-HSV-1 ICP0 antibody Abcam, ab6513, Cambridge, UK), and incubated for 1 h at 37 °C. After wash with PBS, FITC-labeled anti-mouse antibody (Rabbit Anti-Mouse IgG H&L (FITC) Abcam, ab6724, Cambridge, UK) was added, and incubated for 30 min at 37 °C. Infected and uninfected Vero cells were used as controls. The results were recorded using an AxioScop fluorescence microscope (Zeiss, Oberkochen, Germany).
Rabbit anti mouse igg h l
Manufactured by Abcam
Sourced in United Kingdom, United States
Rabbit anti-mouse IgG H&L is a secondary antibody used for the detection and visualization of mouse immunoglobulins in various immunoassays. It is produced in rabbits and recognizes the heavy and light chains of mouse IgG.
Automatically generated - may contain errors
Lab products found in correlation
Bca kit, by Thermo Fisher Scientific (3 mentions)
Goat anti rabbit igg h l, by Abcam (2 mentions)
Ecl system, by Thermo Fisher Scientific (2 mentions)
Ripa lysate, by Beyotime (2 mentions)
Ab6513, by Abcam (1 mentions)
C0265, by Beyotime (1 mentions)
N storm, by Nikon (1 mentions)
Collagen 2 monoclonal antibody, by Thermo Fisher Scientific (1 mentions)
P0202, by Beyotime (1 mentions)
Polyvinylidene fluoride membrane, by Merck Group (1 mentions)
Ecl substrate kit, by Abcam (1 mentions)
Mouse monoclonal anti gapdh, by Abcam (1 mentions)
Chemidoc imaging system, by Bio-Rad (1 mentions)
Lysis buffer, by Beyotime (1 mentions)
Rabbit polyclonal anti v5, by Abcam (1 mentions)
Goat anti mouse hrp, by Thermo Fisher Scientific (1 mentions)
Goat anti rabbit hrp, by Thermo Fisher Scientific (1 mentions)
Rabbit polyclonal anti hif 2α, by Novus Biologicals (1 mentions)
Mouse monoclonal anti hif 1β, by Novus Biologicals (1 mentions)
Mouse monoclonal anti v5 tag, by Thermo Fisher Scientific (1 mentions)
11 protocols using rabbit anti mouse igg h l
1
Visualizing ICP0 Protein in HSV-1-Infected Cells
2
Collagen-II Expression in HNPCs
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In the light of a previous delineation, the expression of Collagen-II was observed through ICC [30 (link)]. The treated HNPCs were collected and washed with PBS, and then fixed with 4% paraformaldehyde at room temperature for 15 min. Next, cells were treated with 0.2% of Triton X-100 (P0096, Beyotime, Shanghai, China) for 10 min, and incubated firstly with 10% goat serum (C0265, Beyotime, Shanghai, China) for 20 min, and then with the diluted primary antibody (Collagen II Monoclonal Antibody; MA1-37,493, Thermo Fisher Scientific, Waltham, Massachusetts, USA) at 4°C overnight. Subsequently, cells were incubated with the diluted secondary antibody (Rabbit Anti-Mouse IgG H&L; ab6728, Abcam, Cambridge, UK) at room temperature for 1 h. Later, the cells were treated with DAB working solution (P0202, Beyotime, Shanghai, China), and then washed with distilled water. Lastly, the Collagen-II content of cells was observed (under 200 × magnification) using fluorescence microscope (N-STORM, Nikon, Tokyo, Japan).
3
Protein Expression Analysis in Kidney Tissues
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Total protein was extracted from kidney tissues using the lysis buffer (Beyotime, China) and quantified using the BCA kit (Thermo Fisher Scientific, MA, USA). Proteins were separated on 10% SDS-PAGE and then transferred onto polyvinylidene fluoride membranes (Millipore, MA, USA). After blocking with 5% skim milk for 1 h, membranes were incubated with primary antibodies at 4°C overnight. Then, membranes were incubated with horseradish peroxidase (HRP)-conjugated secondary antibodies for 1 h under dark condition. Protein bands were visualized using the ECL Substrate Kit (Abcam, UK) and photographed under the ChemiDoc Imaging System (Bio-Rad, CA, USA). The primary antibodies used in Western blotting were rabbit monoclonal anti-PTGS2 (1 : 1,000; GST, MA, USA), mouse monoclonal anti-IL-6 (1 : 500; Abcam, UK), rabbit monoclonal anti-MAPK1 (1 : 2,000; GST), rabbit polyclonal anti-Akt (1 : 1,000; Abcam), rabbit polyclonal anti-p-Akt (1 : 500; Abcam), and mouse monoclonal anti-GAPDH (1 : 500; Abcam). The secondary antibodies were rabbit anti-mouse IgG H&L and goat anti-rabbit IgG H&L (1 : 2,000; Abcam).
4
Antibody validation for HIF-1/2α ChIP-seq and Western blot
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The following antibodies were used for ChIP-seq: rabbit polyclonal anti-HIF-2α (Novus Biologicals, Centennial, CO, #NB100-122), mouse monoclonal anti-HIF-1β (Novus Biologicals, #NB100-124), rabbit polyclonal anti-V5 (Abcam, Cambridge, UK, #ab9116). The following antibodies were used for Cut&Run: mouse monoclonal anti-V5 tag (Thermo Fisher, #R960-25) diluted to 0.01 mg/ml, mouse IgG (Jackson ImmunoResearch #015-000-003) diluted to 0.01 mg/ml, rabbit anti-mouse IgG H&L (Abcam, #ab46540) diluted to 0.01 mg/ml. The following antibodies were used for western blotting: rabbit monoclonal anti-HIF-2α (Cell Signaling, Danvers, MA, #D9E3) diluted at 1:1000, rabbit monoclonal anti-HIF-1β (Cell Signaling, #D28F3) diluted at 1:1000, mouse monoclonal anti-V5 tag (Thermo Fisher, #R960-25) diluted at 1:2500, mouse monoclonal anti-HaloTag (Promega, Madison, WI, #G9211) diluted at 1:1000, mouse monoclonal anti-TBP (Abcam, #ab51841) diluted at 1:2500, goat-anti-mouse-HRP (Thermo Fisher, #31430) diluted at 1:2000, goat-anti-rabbit-HRP (Thermo Fisher, #31462) diluted at 1:2000.
5
Quantifying Epithelial-Mesenchymal Transition in Ovarian Cancer
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Total protein of ovarian cancer tissues, and SKOV3 and CAOV-3 cells, were extracted using radioimmunoprecipitation assay (RIPA) lysate (Beyotime, Shanghai, China). After protein quantification using the bicinchoninic acid method (Waltham, MA, USA), 80 μg protein was subject to sodium dodecyl sulfate-polyacrylamide gel electrophoresis for separation, and was then transferred onto polyvinylidene difluoride membranes. After that, the membranes were blocked (1 h) in TBS-T buffer containing 5% skim milk. Next, the membranes were subject to an overnight incubation (4°C) with the following primary antibodies (all from Abcam, Cambridge, MA, USA): anti-E Cadherin antibody (Cat# ab40772, 1/10000), anti-N Cadherin antibody (Cat# ab18203, 1/1000), anti-SNAIL antibody (Cat# ab53519, 1/1000), anti-Vimentin antibody (Cat# ab8978, 1/1000), and anti-3-phosphate glyceraldehyde dehydrogenase (GAPDH) antibody (Cat# ab8245, 1/500). After three Tris-Buffered Saline Tween-20 (TBST) washes, the membranes were hybridized (room temperature, 1.5 h) with the horseradish peroxidase-linked secondary antibody rabbit anti-mouse IgG H&L (Cat# ab6728, 1/2000, Abcam). Signal detection was performed using the enhanced chemiluminescence (ECL) system (Life Technologies Corporation); the relative protein levels were calculated by normalization to GAPDH.
6
Rapid Malaria Antigen Detection Assay
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A 25-mm-wide nitrocellulose membrane (GE Healthcare, IL) was cut into 300-mm-long strips using a guillotine cutter. A solution of 200 μg/mL rabbit anti-mouse IgG H&L (Abcam, UK) and 800 µg/mL mouse anti-Plasmodium falciparum HRP2 IgM (ICL, Inc., OR) were dispensed on nitrocellulose membrane strips as control and test lines, respectively, using an automated liquid dispensing platform (BioDot, CA). The nitrocellulose membrane card was incubated at 37 °C for 2 h and cut into 3-mm-wide strips.
7
Rapid Influenza A Detection Assay
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Rabbit anti-mouse IgG H&L (horseradish peroxidase (HRP)) and goat anti-mouse IgG H&L (FICT) ab6758 were bought from Abcam (Cambridge, UK). Europium nanoparticles (200 nm diameter) were purchased from Bangs Laboratories Inc. (Fishers, IN, USA). Colloidal gold (40 nm diameter) was obtained from Bore Da Biotech Co., (Gyeonggi, Republic of Korea). N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysulfosuccinimide sodium salt (Sulfo-NHS) were acquired from Thermo Scientific (Waltham, MA, USA). All other chemicals were purchased from Sigma-Aldrich (St. Louis, MO, USA) and used without further purification. Anti-influenza A nucleoprotein (NP) (Clone 3D3) was provided by Professor Ho-Joon Shin, Ajou University, Suwon, Republic of Korea.
8
Apoptosis Pathway Analysis in Cell Culture
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The PVDF membranes (catalog no. IPVH 20200, Millipore), mouse monoclonal β-actin (A2228) Horseradish peroxidase conjugated anti-rabbit IgG (A1949), STZ (S0130) were purchased from Sigma-Aldrich, St Louis, USA. Rabbit polyclonal Bcl-2 (N-19): SC492, rabbit polyclonal Bax (SC- 6236), were purchased from Santa Cruz Biotechnology, Inc. Cleaved PARP-1 ([E51], ab32064), rabbit anti-mouse IgG H&L (conjugated with FITC-green fluorescence, ab6724)and goat anti-rabbit IgG H&L (conjugated with TRITC-red, ab6718) were purchased from Abcam, USA. Active Caspase-3 (Asp175) purchased from Cell Signaling Technology, Inc. Prestained protein molecular weight marker was obtained from Hi-media Pvt. Ltd., Kolkata, India. All biochemical were of analytical grade. Biochemical kits were purchased from Accurex Biomedical Pvt. Ltd., Biosar, Thane.
9
CUT&Tag for H3K27ac profiling
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Cleavage under targets and tagmentation (CUT&Tag) was conducted as previously described (26 (link)). A total of 1 × 105 cells were incubated in the antibody buffer with 10 μl pre-cleaned A-MyOne T1 beads (Thermo Fisher Scientific, #65601) and 1 μl H3K27ac antibody (Active Motif, #91193). After primary antibody binding, cells were incubated with 1 μl secondary antibody (Rabbit Anti-Mouse IgG H&L, Abcam, #ab6709) in Dig-wash buffer for 1 h at room temperature and washed using a Dig-wash buffer five times. Then, adapters were added to the fragments with the pA-Tn5 Kit (EpiCypher, #15-1017), and the mixture was washed twice using a Dig-wash buffer. The beads were resuspended in tagmentation buffer and incubated for 1 h at 37°C. After DNA extraction, library was constructed using KAPA HiFi HotStart ReadyMix (Roche, #07958927001), and sequenced by NovaSeq 6000.
10
Western Blot Analysis of Cellular Proteins
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First of all, proteins were extracted from cells by using RIPA buffer with 1% PMSF and the concentration of protein is evaluated by BCA assay. And then, the proteins were segregated by SDS‐PAGE and electrophoretically transferred onto PVDF membranes (Millipore, Billerica, MA). Then, the membranes were blocked with bovine serum albumin (BSA) (Sigma‐Aldrich, St. Louis, MO) and treated with specific antibodies. GAPDH protein is the loading control. The primary antibodies were as following: Bcl‐2 (#4223), Bax (#2772), CDK4 (#12790), Cyclin D1 (#2978) (above antibodies were from Cell Signaling Technology, Inc, Danvers, MA, USA), GAPDH (ab8245, Abcam) and CHD1 (20576‐1‐AP, Protein‐Tech). The secondary antibody was Rabbit Anti‐Mouse IgG H&L (ab6728, Abcam). Next, ECL chemiluminescent detection system (Thermo Fisher Scientific, Rochester, NY) was applied to visualize protein bands. In the end, all of the proteins were exposed to X‐ray film. All experimental steps were performed for at least three times.
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