The following antibodies were used: anti-SKI (sc-9140, Santa Cruz); anti-RUNX1 (ab23980, Abcam); anti-CDK2 (sc-163, Santa Cruz); anti-Flag (F3163, Sigma); anti-HA (HA.11 clone 16B12; Covance); anti-H3K4me3 (07-473, Millipore); anti-H3K27ac (39133, Active Motif); rabbit IgG (I5006, Sigma).
Anti ha ha 11 clone 16b12
Manufactured by Fortrea
Sourced in United States
Anti-HA (HA.11 clone 16B12) is a monoclonal antibody that specifically binds to the Hemagglutinin (HA) epitope tag. It can be used for the detection and purification of HA-tagged proteins.
Automatically generated - may contain errors
Lab products found in correlation
Ab23980, by Abcam (1 mentions)
Anti h3k4me3 07 473, by Merck Group (1 mentions)
Anti h3k27ac 39133, by Active Motif (1 mentions)
Anti cdk2 sc 163, by Santa Cruz Biotechnology (1 mentions)
Rabbit igg i5006, by Merck Group (1 mentions)
F1804, by Merck Group (1 mentions)
Ab70472, by Abcam (1 mentions)
Co2 independent medium, by Thermo Fisher Scientific (1 mentions)
Pegfp n1, by Takara Bio (1 mentions)
Infrared dye coupled secondary antibodies, by LI COR (1 mentions)
Odyssey infrared imaging system, by LI COR (1 mentions)
Anti tsg101 c 2, by Santa Cruz Biotechnology (1 mentions)
Tween 20, by Thermo Fisher Scientific (1 mentions)
Anti v5, by Thermo Fisher Scientific (1 mentions)
Anti flag, by Merck Group (1 mentions)
Horseradish peroxidase conjugated species specific secondary antibodies, by Dianova (1 mentions)
Las 4000 imaging system, by Bio-Rad (1 mentions)
Western lightning chemiluminescence reagent plus, by PerkinElmer (1 mentions)
Anti gst, by GE Healthcare (1 mentions)
Las 4000, by Cytiva (1 mentions)
6 protocols using anti ha ha 11 clone 16b12
1
Antibody Validation for Protein Analysis
2
Whole Cell Extract Preparation and Analysis
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Whole cell extract preparation, western blotting and co-immunoprecipitation experiments were performed as described previously [5 (link),6 (link)]. Antibodies used were: anti-BrdU (ab12219, abcam), anti-FLAG (F1804, Sigma), anti-HA (HA.11 clone 16B12, Covance), anti-PCNA (ab70472, abcam).
3
HIV-1 Mutant Analysis and Protein Detection
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HIV-1ΔR8.2 (HIV-1NL4-3 R9ΔApa [70 (link)]) and HIV R9 were used. Its late domain mutants, ΔPTAP and ΔYP were previously described [43 ]. Humanized Gag was produced as previously described [71 (link)]. ALIX (NM_013374), Tsg101 (NM_006292), CHMP4b (NM_176812) and VPS4A (NM_013245) were kindly provided by Dr. Wesley Sundquist (university of Utah) and were all HA N-terminally tagged. GFP ORF was cloned from peGFP-N1 (Clontech). Point mutations were introduced using the Quick Change site directed mutagenesis kit (Stratagene).
All cell lines used were grown in complete DMEM medium under standard conditions, excepted for TIRF experiments where cells were incubated in CO2-independent medium (LifeTechnologies).
Anti-ALIX [44 (link)], anti-Tsg101 (C-2, Santa Cruz Biotech.), anti-HA (HA.11 clone 16B12, Covance), anti-p24 (183-H12-5C, NIH AIDS Reagent Program), anti-p17 (17–1, Santa Cruz Biotech.), anti-RT (MAb21, NIH AIDS Reagent Program), anti-PR (1696, Santa Cruz Biotech.), and infrared dye coupled secondary antibodies (LI-COR) were used for immunoprobing. Scanning was performed with the Odyssey infrared imaging system (LI-COR) in accordance with the manufacturer’s instructions at 700 or 800 nm, accordingly.
All cell lines used were grown in complete DMEM medium under standard conditions, excepted for TIRF experiments where cells were incubated in CO2-independent medium (LifeTechnologies).
Anti-ALIX [44 (link)], anti-Tsg101 (C-2, Santa Cruz Biotech.), anti-HA (HA.11 clone 16B12, Covance), anti-p24 (183-H12-5C, NIH AIDS Reagent Program), anti-p17 (17–1, Santa Cruz Biotech.), anti-RT (MAb21, NIH AIDS Reagent Program), anti-PR (1696, Santa Cruz Biotech.), and infrared dye coupled secondary antibodies (LI-COR) were used for immunoprobing. Scanning was performed with the Odyssey infrared imaging system (LI-COR) in accordance with the manufacturer’s instructions at 700 or 800 nm, accordingly.
4
Western Blot Analysis of HCV Proteins
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Proteins were separated in polyacrylamide-Tricine gels. After SDS-PAGE, proteins were transferred onto a nitrocellulose membrane (Pall, USA). The membrane was blocked with 5% (w/v) dried skim milk in phosphate-buffered saline with 0.05% (v/v) Tween 20 (Invitrogen). For antigen detection, anti-NS5A 9E10 [60 (link)], mouse monoclonal antibody against NS3 of the JFH-1 isolate (4D11) (generated in a cooperation between Harish Ramanathan, Michael Engle, Michael S. Diamond and Brett D. Lindenbach) or anti-NS3 (2E3) [61 (link)], anti-NS2 (YAL-4-70-8, Cell Essentials), anti-NS4B [62 (link)], anti-FLAG (Sigma), anti-V5 (Invitrogen), anti-HA (HA.11 clone 16B12, Covance) and anti-GST (GE Healthcare), antibodies were used in 2% (w/v) dried skim milk in phosphate-buffered saline with 0.05% (v/v) Tween 20. For primary antibody detection, horseradish peroxidase-conjugated species-specific secondary antibodies (Dianova) were used at a 1:3000 dilution and Western Lightning Chemiluminescence Reagent Plus (Perkin Elmer) was applied prior to imaging using a LAS 4000 imaging system (Biorad, Munich). Quantifications of Western blots were carried out using ImageJ 1.47t software (NIH, Bethesda).
5
Rtt106 Chromatin Immunoprecipitation in Yeast
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Yeast strains were grown to an OD600 of 0.25 in YPD. Alpha factor was added to arrest cells in G1 and released into YPD containing 0.2M hydroxyurea at 23°C for 60 minutes. Formaldehyde (1% final concentration) was added and incubated with rotation first at room temperature for 20 minutes and then at 4°C overnight. Cells were washed 3 times with ice-cold 1X Phosphate buffered saline. Cells were pelleted and frozen at -80°C. Rtt106 ChIP using anti-HA (HA.11 clone 16B12, Covance) and data analysis were performed as described previously [6 (link)].
ChIP-Seq data are uploaded to Array Express under accession number: E-MTAB-6985
ChIP-Seq data are uploaded to Array Express under accession number: E-MTAB-6985
6
Antibody-Based SARS-CoV-2 Antigen Detection
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For antigen detection, mouse monoclonal antibody 8.12.7 against NS3/NS2-3 [61 (link)], anti-E2 (SCR48 6.6.11) (kindly provided by H.-J. Thiel, [Justus-Liebig University, Gießen]) [62 (link)], anti-NS4A (GH4A1 [4B7]), anti-NS5A (GLBVD5A1 [11C]), anti-NS5B (GLBVD5B1 [9A]) (kindly provided by T. Rümenapf and B. Lamp [University of Veterinary Medicine, Vienna, Austria]) [43 (link)], anti-HA (HA.11 clone 16B12, Covance, New Jersey, USA), anti-GST (New England Biolabs), anti-myc (New England Biolabs) were used. Species specific Cyanogen-3-labeled (Cy3) or peroxidase-coupled (PO) antibodies were obtained from Dianova (Hamburg). Quantitative Western blot analyses were performed using secondary antibody coupled to IRdye-800 obtained from LI-COR Biosciences (Lincoln, Nebraska, USA).
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