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5
reviewed by: Anonymous
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WB |
Mouse |
05/08/2020 |
Summary
Application | Western Blot | Sample Tested | 3T3-L1 mouse embryonic fibroblast adipose-like cell line | Species | Mouse | Lot | lot A3 |
Comments
Comments | skimmed milk 5% primary : 1:2000 secondary: 1: 200000 |
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4
reviewed by: Anonymous
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IHC-P |
Mouse |
06/07/2019 |
Summary
Application | Immunohistochemistry-Paraffin | Sample Tested | PDX mouse | Species | Mouse | Lot | Lot A3 |
Comments
Comments | Working Dilution 1:100, EDTA pH 9.0; detection kit Polymer-HRP. |
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4
reviewed by: an shiyu
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WB |
Goat |
05/15/2018 |
Summary
Application | Western Blot | Sample Tested | laydig cell | Species | Goat | Lot | NBP1-04676SS |
Comments
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4
reviewed by: Anonymous
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WB |
Mouse |
07/17/2017 |
Summary
Application | Western Blot | Sample Tested | BMDM | Species | Mouse | Lot | A5 |
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4
reviewed by: Anonymous
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IP |
Rat |
04/24/2017 |
Summary
Application | Immunoprecipitation | Sample Tested | PC-12 rat adrenal pheochromocytoma cell line | Species | Rat | Lot | A6 |
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4
reviewed by: Anonymous
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IF |
Mouse |
01/12/2016 |
Summary
Application | Immunofluorescence | Sample Tested | primary bone marrow derived macrophages | Species | Mouse |
Blocking
Details
Comments
Comments | Murine primary bone marrow derived macrophages probed with rabbit HIF1a antibody (1:100) in blocking buffer at 4°C overnight, secondary goat anti-rabbit AF647 (1:250) for 2 h at ambient temperature, Cells were mounted with Vectashield containing DAPI for imaging. Image (top right) derived from merge of DAPI and AF647 staining. White arrows point at cells containing nuclear HIF1a. Unstained and secondary antibody controls shown in bottom panels, Magnification, 63X |
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4
reviewed by: Anonymous
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WB |
Human |
09/07/2015 |
Summary
Application | Western Blot | Sample Tested | Human cancer cell whole cell lysate | Species | Human |
Blocking
Blocking Details | Blocking with 5% non-fat milk for 30 min |
Primary Anitbody
Dilution Ratio | 1:1000, overnight |
Secondary Antibody
Secondary Description | Donkey anti-rabbit IgG, HRP | Secondary Concentration | 1:2000 |
Details
Detection Notes | Regular Protocol |
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4
reviewed by: Zachary Patinkin
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Simple Western |
Human |
06/09/2015 |
Summary
Application | Simple Western | Sample Tested | Human mesenchymal stem cell whole cell lysate | Species | Human | Lot | A5 |
Blocking
Blocking Details | Protein Simple Antibody Diluent, 5 min, 25C |
Primary Anitbody
Dilution Ratio | 1:200, 30 minutes, Protein Simple Antibody Diluent |
Secondary Antibody
Secondary Description | Protein Simple Anti-Rabbit Secondary | Secondary Manufacturer Cat# | 042-206 | Secondary Concentration | undiluted |
Details
Detection Notes | WES multi-image analysis |
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4
reviewed by: Enrica Borsi
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WB |
Human |
01/09/2015 |
Summary
Application | Western Blot | Sample Tested | Myeloma cells whole cell lysate | Species | Human |
Blocking
Blocking Details | Blocking buffer Milk 5% |
Details
Detection Notes | Western blot |
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4
reviewed by: Anonymous
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WB |
Human |
12/12/2014 |
Summary
Application | Western Blot | Sample Tested | See PMID 22923663 | Species | Human |
Blocking
Blocking Details | See PMID 22923663 |
Details
Detection Notes | See PMID 22923663 |
Comments
Comments | Published in PMID: 22923663 |
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4
reviewed by: Ming Lu
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WB |
Human |
08/15/2014 |
Summary
Application | Western Blot | Sample Tested | Human pancreatic cancer cell lysates | Species | Human | Lot | A4 | Comments | Although I got my result as expected, this antibody has weak bands and dirty background in my western blotting. |
Blocking
Blocking Details | 2% cold fish geletin in TBS buffer, room temperature for 1hour |
Primary Anitbody
Dilution Ratio | 1:1000 overnight at 4C in blocking buffer |
Secondary Antibody
Secondary Description | Goat anti-rabbit IgG, HRP | Secondary Manufacturer Cat# | Cell Signaling, #7074 | Secondary Concentration | 1:5000 |
Details
Detection Notes | 4 min exposure shows band at correct MW. |
Comments
Comments | Although I got my result as expected, this antibody has weak bands and dirty background in my western blotting. |
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5
reviewed by: Xiuquan Luo
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WB |
Human |
07/18/2014 |
Summary
Application | Western Blot | Sample Tested | Caki-1 cell lysate | Species | Human |
Blocking
Blocking Details | 1xPBST with 5% non-fat milk, RT 1 hour |
Primary Anitbody
Dilution Ratio | 1xpBST with 5% non-fat milk, overnight 4C, 1:1000 dilution |
Secondary Antibody
Secondary Description | Goat anti-rabbit IgG-HRP | Secondary Manufacturer Cat# | sc-2004 | Secondary Concentration | 1:2000 |
Details
Detection Notes | ECL, exposured for 1 min |
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4
reviewed by: Anonymous
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WB |
Mouse |
07/05/2014 |
Summary
Application | Western Blot | Sample Tested | large intestinal lamina propria leukocytes (LPLs). | Species | Mouse |
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5
reviewed by: Paramananda Saikia
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WB |
Mouse |
06/30/2014 |
Summary
Application | Western Blot | Sample Tested | Mouse | Species | Mouse | Lot | A4 |
Blocking
Primary Anitbody
Secondary Antibody
Secondary Description | goat anti-rabbit |
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5
reviewed by: Alshaimaa Adawy
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WB |
Human |
04/29/2014 |
Summary
Application | Western Blot | Sample Tested | nuclear fractions from T47-D beast cancer cell line | Species | Human | Lot | A5 |
Blocking
Blocking Details | blocking: 5% BSA for 1 hour at room temprature |
Primary Anitbody
Dilution Ratio | 1:1000, overnight at 4 C in BSA |
Secondary Antibody
Secondary Description | Anti-rabbit IgG, HRP-linked Antibody | Secondary Manufacturer Cat# | 7074s, cell signaling | Secondary Concentration | 1:5000 |
Details
Detection Notes | Vilber Lourmat imager, using home made ECL, exposure time 3 minutes |
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4
reviewed by: Anonymous
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WB |
Other |
03/14/2014 |
Summary
Application | Western Blot | Species | Other | Lot | A4 | Pub Med ID | 21193390 | File | View PDF |
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4
reviewed by: Lidan Zhao
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WB |
Other |
08/26/2013 |
Summary
Application | Western Blot | Sample Tested | pig satellite cell nuclear protein | Species | Other | Comments | I first ordered NB100-105 and tried several times, but it didn't work with my samples. I contacted technical support and they sent me a free vial of NB100-449 as a replacement. I found that this antibody worked well both on the positive control and my pig samples. |
Blocking
Blocking Details | 5% milk in TBST |
Primary Anitbody
Dilution Ratio | 1:500 in 5% milk, 4 C overnight |
Secondary Antibody
Secondary Description | IRDye 800CW Goat anti-Rabbit IgG | Secondary Concentration | 1:20000 |
Details
Detection Notes | IRDye conjugated secondary scanned by Licor Odyssey system. Used Novus COS-7 nuclear extracts (NB800-PC26) as positive control |
Comments
Comments | I first ordered NB100-105 and tried several times, but it didn't work with my samples. I contacted technical support and they sent me a free vial of NB100-449 as a replacement. I found that this antibody worked well both on the positive control and my pig samples. |
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5
reviewed by: Anonymous
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WB |
Human |
11/05/2012 |
Summary
Application | Western Blot | Sample Tested | human cell lysates | Species | Human |
Blocking
Blocking Details | 5% Non fat milk |
Primary Anitbody
Secondary Antibody
Secondary Description | anti-rabbit-HRP | Secondary Manufacturer Cat# | Sigma | Secondary Concentration | 1:1000 |
Details
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5
reviewed by: Annie Curtis
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WB |
Mouse |
08/24/2012 |
Summary
Application | Western Blot | Sample Tested | mouse peritoneal and bone marrow derived macrophages | Species | Mouse | Lot | A4 | Comments | We tried numerous HIF1alpha antibodies with little success, we finally got this one to work but one thing that is necessary is that you need to lyse your cells directly in loading buffer. We used about 100uL of Laemmli buffer per 6 well dish, then boil immediately after lysing. Samples can then be either loaded directly onto the gel or stored at -20. However, we did find that if samples were stored for too long at -20 then we did lose the ability to detect the protein. |
Blocking
Blocking Details | blocked in 5% milk for 1 hour at room temperature |
Primary Anitbody
Dilution Ratio | 1:1000 overnight at 4deg C, it has to be an overnight incubation to work |
Secondary Antibody
Secondary Description | Goat anti-rabbit IgG, HRP | Secondary Concentration | 1: 1000 |
Details
Detection Notes | ECL exposure of 2 minutes needed |
Comments
Comments | We tried numerous HIF1alpha antibodies with little success, we finally got this one to work but one thing that is necessary is that you need to lyse your cells directly in loading buffer. We used about 100uL of Laemmli buffer per 6 well dish, then boil immediately after lysing. Samples can then be either loaded directly onto the gel or stored at -20. However, we did find that if samples were stored for too long at -20 then we did lose the ability to detect the protein. |
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5
reviewed by: kimberley tam
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WB |
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08/07/2012 |
Summary
Application | Western Blot | Lot | A2 | Special Applications | Excellent antibody. Tested with mouse | Pub Med ID | 20600586 |
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4
reviewed by: Anonymous
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WB |
|
05/09/2012 |
Summary
Application | Western Blot | Pub Med ID | 16611993 | File | View PDF |
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4
reviewed by: Anonymous
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WB |
Rat |
01/03/2012 |
Summary
Application | Western Blot | Sample Tested | PC12 cell whole lysate, Sample Amount: 25ug | Species | Rat | Lot | A4 | Comments | This antibody could detect HIF-1a but weak. |
Blocking
Blocking Details | Blocking Buffer: 5% Milk in TBST, Blocking Time: 1 hour, Blocking Temp: Room temperature |
Primary Anitbody
Dilution Ratio | Dilution Ratio: 1:1000, Incubation Dilution Buffer: blocking buffer, Incubation Time: overnight, Incubation Temp: 4 C |
Secondary Antibody
Secondary Description | Secondary Ab: anti-Rabbit IgG, Secondary Ab Dilution Ratio: 1:5000 |
Details
Detection Notes | Detection Method: ECL Plus |
Comments
Comments | This antibody could detect HIF-1a but weak. |
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4
reviewed by: Anonymous
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WB |
Human |
06/08/2011 |
Summary
Application | Western Blot | Sample Tested | HUVEC whole cell lysate, Sample Amount: 20 microg | Species | Human | Comments | Western blot analysis showing that the increase in HIF-1alpha levels triggered by hypoxia in HUVEC is prevented by a specific siRNA against HIF-1alpha (siHIFa).(N: Normoxia, H: Hypoxia; siRAND: Random siRNA | Pub Med ID | 21622104 | File | View PDF |
Blocking
Blocking Details | Blocking Buffer: 5%Milk in TBST, Blocking Time: overnight, Blocking Temp: 4 degrees Celsius |
Primary Anitbody
Dilution Ratio | Dilution Ratio: 1:500, Incubation Dilution Buffer: 5% Milk in TBST, Incubation Time: 1 hour, Incubation Temp: room temperature |
Secondary Antibody
Secondary Description | Secondary Ab: anti-rabbit DAKO, Secondary Ab Dilution Ratio: 1:2000 | Secondary Manufacturer Cat# | P0448 |
Details
Detection Notes | Method: SuperSignal West, Time: 3min, Controls: Hypoxia/normoxia in random transfected cells, Bands: Multiband at 110kDa |
Comments
Comments | Western blot analysis showing that the increase in HIF-1alpha levels triggered by hypoxia in HUVEC is prevented by a specific siRNA against HIF-1alpha (siHIFa).(N: Normoxia, H: Hypoxia; siRAND: Random siRNA |
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4
reviewed by: Anonymous
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WB |
Mouse |
12/22/2010 |
Summary
Application | Western Blot | Sample Tested | cerebellar neurons | Species | Mouse | Comments | This Hif1 alpha antibody is the only one we have found to work consistently well. We use it 1:500 in 1% BSA and reuse it for 4-5 times. We get a a doublet, the expected band at around 100KDa and another band at about 130KDa or so (right below the 150KDa marker.) |
Blocking
Blocking Details | Blocking Buffer: 3% BSA in TBST, Blocking Time: 1 hour, Blocking Temp: Room temperature |
Primary Anitbody
Dilution Ratio | Primary Ab Dilution Ratio: 1:500, Primary Ab Incubation Time: overnight, Primary Ab Incubation Temp: 4 degrees Celsius |
Secondary Antibody
Secondary Description | Secondary Ab Dilution Ratio: 1:5000 |
Details
Detection Notes | Detection Method: ECL, Exposure Time: 5 minutes |
Comments
Comments | This Hif1 alpha antibody is the only one we have found to work consistently well. We use it 1:500 in 1% BSA and reuse it for 4-5 times. We get a a doublet, the expected band at around 100KDa and another band at about 130KDa or so (right below the 150KDa marker.) |
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4
reviewed by: William Lin
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IP |
Human |
12/03/2010 |
Summary
Application | Immunoprecipitation | Sample Tested | HEK 293 | Species | Human | Lot | A3 | Comments | I used BD mouse anti-human HIF1 antibody for WB because I didn't want to detect the rabbit IgG from the NB100-449 antibody during the IP. Lysate Amount: 1mg, Lysate Preclear: agarose A, Diluted Lysate Concentration: 2ug/ul, Ab Dilution Ratio: 1:100, Protein Capture: agarose A, Collection Method: 4000 rpm centrifugation, Protein Reduction: 2X sample buffer, with SDS/bME, Bead Removal: 4min, 4C |
Blocking
Blocking Details | Blocking Buffer: 5% milk in TBST, Blocking Time: 2 hours, Blocking Temp: 4 degrees Celsius |
Primary Anitbody
Dilution Ratio | Incubation Dilution Buffer: 5% milk/TBST, Dilution Ratio: 1:2000, Incubation Time: overnight, Incubation Temp: 4 C |
Secondary Antibody
Secondary Description | Secondary Ab: anti-human IgG-HRP, Secondary Ab Dilution: 1:10000 | Secondary Manufacturer Cat# | promega |
Details
Detection Notes | Method: ECL, Exposure Time: 2 min, Pos.Ctrl: DFO treated, Neg.Ctrl: no treatment, Specific Bands: 116kDa |
Comments
Comments | I used BD mouse anti-human HIF1 antibody for WB because I didn't want to detect the rabbit IgG from the NB100-449 antibody during the IP. Lysate Amount: 1mg, Lysate Preclear: agarose A, Diluted Lysate Concentration: 2ug/ul, Ab Dilution Ratio: 1:100, Protein Capture: agarose A, Collection Method: 4000 rpm centrifugation, Protein Reduction: 2X sample buffer, with SDS/bME, Bead Removal: 4min, 4C |
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5
reviewed by: Anonymous
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WB |
Other |
03/11/2010 |
Summary
Application | Western Blot | Sample Tested | Colon Cancer Cell lines, Sample Amount: 50ug | Species | Other | Lot | A2 | Comments | The lanes on the left are HCT116 treated with increasing concentrations of DMOG. The lanes on the right are HCT116 Hif1-/- cells with the same treatment. |
Blocking
Blocking Details | Blocking Buffer: 5% Blotto, Blocking Time: 1 hour, Blocking Temp: Room temperature |
Primary Anitbody
Dilution Ratio | Dilution Ratio: 1/5000, Incubation Dilution Buffer: 5% BSA in TNS with Tween, Incubation Time: overnight, Incubation Temp: 4C |
Secondary Antibody
Secondary Description | Secondary Ab: Cell Signaling Goat Anti-Rabbit HRP, Secondary Ab Dilution Ratio: 1/10000 |
Details
Detection Notes | Detection Method: ECL Plus, Specific Bands: 120 kDa |
Comments
Comments | The lanes on the left are HCT116 treated with increasing concentrations of DMOG. The lanes on the right are HCT116 Hif1-/- cells with the same treatment. |
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3
reviewed by: Anonymous
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WB |
Mouse |
12/21/2009 |
Summary
Application | Western Blot | Sample Tested | human tumor model mouse, Sample Amount: 12ug | Species | Mouse | Comments | Nuclear protein extracts: Chen et al. J Cell Biochem 2001;82(3):512-521. |
Blocking
Blocking Details | Blocking Buffer: 5% BSA, Blocking Time: 1 hour, Blocking Temp: Room temperature |
Primary Anitbody
Dilution Ratio | Dilution Ratio: 1/500, Incubation Dilution Buffer: TBST with 5% milk, Incubation Time: overnight, Incubation Temp: 4 C |
Secondary Antibody
Secondary Description | Secondary Ab: Anti-rabbit IgG HRP-linked, Cell Signaling #7047, Secondary Ab Dilution Ratio: 1/3000 |
Details
Detection Notes | Detection Method: ECL, Exposure Time: 5 - 10 min., Specific Bands: 100 kDa |
Comments
Comments | Nuclear protein extracts: Chen et al. J Cell Biochem 2001;82(3):512-521. |
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5
reviewed by: Anonymous
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WB |
Mouse |
04/08/2009 |
Summary
Application | Western Blot | Sample Tested | Mouse Neuroblastoma, Sample Amount: 25ug | Species | Mouse | Lot | A2 | Comments | "Load: Empty Vector Normoxia Empty Vector Hypoxia</p><p>Hif1a over expression Normoxia Hif1a over expression Hypoxia" |
Blocking
Blocking Details | Blocking Buffer: 5% NFDM with TBS and.5% Tween 20, Blocking Time: 90 minutes, Blocking Temp: 25 degrees Celsius |
Primary Anitbody
Dilution Ratio | Dilution Ratio: 1/500, Incubation Dilution Buffer: 5% NFDM w/ TBS and.5% Tween 20, Incubation Time/Temp: overnight(16hrs) at 4C |
Secondary Antibody
Secondary Description | Secondary Ab: Santa Cruz Goat anti Rabbit Secondary, Secondary Ab Dilution Ratio: 1/5000 |
Details
Detection Notes | Method: Chemiluminescence onto Kodak XAR film, Exposure Time: 2-15 min, Bands: 120,60 kDa(specific), 50 kDa(non-specific) |
Comments
Comments | "Load: Empty Vector Normoxia Empty Vector Hypoxia</p><p>Hif1a over expression Normoxia Hif1a over expression Hypoxia" |
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