Antibody against FABP5 was purchased from R&D systems (Minneapolis, MN, USA); anti-p-AKT, anti-p-mTOR, and anti-Ki-67 from Cell Signaling (Danvers, MA, USA); anti-Flag from Sigma-Aldrich (St. Louis, MO, USA); anti-B-tubulin and anti-Lamin B from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-GFP from Thermo Fisher Scientific (Waltham, MA, USA); anti-HA from GeneTex (Irvine, CA, USA). Anti-HIF-1α was generated against human HIF-1α in rabbits, and a monoclonal anti-hydroxylated Asn 803 of HIF-1α was raised in mouse as described previously61 (link),62 (link).
Anti ha
Manufactured by GeneTex
Sourced in United States
The Anti-HA is a laboratory equipment product that is used to detect and identify proteins tagged with the hemagglutinin (HA) epitope. It provides a reliable and specific method for the detection and analysis of HA-tagged proteins in various experimental applications.
Automatically generated - may contain errors
Lab products found in correlation
Anti flag, by Merck Group (3 mentions)
Anti gfp, by GeneTex (2 mentions)
Anti gfp, by Thermo Fisher Scientific (1 mentions)
Anti p akt, by Cell Signaling Technology (1 mentions)
Anti β tubulin, by Santa Cruz Biotechnology (1 mentions)
Anti lamin b, by Santa Cruz Biotechnology (1 mentions)
Anti p mtor, by Cell Signaling Technology (1 mentions)
Anti ki67, by Cell Signaling Technology (1 mentions)
Dynamag magnet, by Thermo Fisher Scientific (1 mentions)
Enhanced chemiluminescence reagent, by GE Healthcare (1 mentions)
Dynabeads protein g, by Thermo Fisher Scientific (1 mentions)
Mab1501, by Merck Group (1 mentions)
Anti ha, by Cell Signaling Technology (1 mentions)
Anti pb1, by GeneTex (1 mentions)
Anti pa, by GeneTex (1 mentions)
Anti m1, by GeneTex (1 mentions)
Anti ns1, by GeneTex (1 mentions)
Anti pb2, by GeneTex (1 mentions)
Anti np, by GeneTex (1 mentions)
Horseradish peroxidase conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
7 protocols using anti ha
1
Antibody Sources for Protein Detection
2
Immunoprecipitation and Immunoblotting of Influenza Viral Proteins
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Cells were harvested in lysis buffer (50 mM Tris pH 8.0, 5 mM NaCl, 0.5% NP-40, and 1X protease inhibitor), frozen and thawed three times, and then the proteins were recovered. Immunoprecipitation (IP) proceeded overnight at 4 °C in IP buffer containing antibodies against Grail or Flag. The IP mixture was then incubated with Dynabeads Protein G (Invitrogen) for 1 h prior to isolation using a DynaMag magnet and washing three times with SNNTE buffer (5% sucrose, 1% NP-40, 0.5 M NaCl, 50 mM Tris pH 7.4, and 5 mM EDTA). The immunoprecipitates were resuspended in SDS-PAGE sample buffer, boiled, and loaded onto a gel. Following separation, the proteins were transferred to a nitrocellulose membrane and the blot was probed with antibodies diluted in PBS/Tween 20 with 5% non-fat milk. Antibody detection was carried out using enhanced chemiluminescence reagents (GE Healthcare), as described by the manufacturer. The primary antibodies used for immunoblotting were: anti-PA (GeneTex), anti-PB1 (GeneTex), anti-PB2 (GeneTex), anti-HA (GeneTex), anti-NA (GeneTex), anti-NP (GeneTex), anti-M1 (GeneTex), anti-M2 (GeneTex), anti-NS1 (GeneTex), anti-HA (81B8, Cell Signaling, USA), anti-beta actin (MAb1501, Chemicon), and anti-Grail antibodies.
3
Western Blot Analysis of Cellular Proteins
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Cells were lysed with RIPA buffer containing a cocktail of protease (Sigma). Protein samples were separated by SDS-PAGE and transferred to a polyvinylidene difluoride (PVDF) transfer membrane (Hybond-P; GE Healthcare). The nonspecific antibody binding sites were blocked with milk in Tris-buffered saline–Tween 20 and then reacted with the primary antibodies anti-HA (GTX29110 [GeneTex] or 3724 [Cell Signaling]), anti-Ubc9 (4786; Cell Signaling), anti-actin (A5441; Sigma), anti-GFP (GTX26556GeneTex), anti-STAT1 (9175; Cell Signaling), anti-STAT2 (05-693; Upstate), and anti-RIG-I (3743; Cell Signaling). Anti-NS5 and -NS3 antibodies are gifts from Huey-Nan Wu. Blots were treated with horseradish peroxidase-conjugated secondary antibody (Thermo Scientific), and signals were detected by enhanced chemiluminescence (ECL; Millipore).
4
Western Blot for Protein Expression
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Cells were washed with ice-cold PBS, harvested in ice-cold PBS, and pelleted by centrifugation at 500 g for 5 minutes. Cell pellets were resuspended with Radioimmunoprecipitation assay (RIPA) buffer (Sigma Aldrich, Saint Louis, MO) and rotated at 4 °C for 1 hour. Lysates were cleared by centrifugation at 16,000 g for 10 minutes and supernatants were analyzed for total protein concentration using a BCA kit (Pierce, Rockford, Illinois). 30 μg of total protein lysate was taken from each sample and fractionated on a 4–12% gradient Novex® Bis-Tris Bolt® SDS-PAGE gel via electrophoresis. Proteins were transferred onto 0.45 μm nitrocellulose membranes and incubated for 1hr in Odyssey® PBS blocking buffer (Li-Cor Biosciences, Lincoln, NE). Membranes were probed with anti-HA (1:5,000, GeneTex, Irvine, CA) and anti-NaK-ATPase (1:2,500, GeneTex, Irvine, CA) primary antibodies in Odyssey® PBS blocking buffer overnight at 4 °C. anti-HA and anti-NaK-ATPase signals were detected using IRdye 800 donkey anti-rabbit (1:20,000, Li-Cor Biosciences, Lincoln, NE) and IRdye 680 donkey anti-mouse (1:20,000, Li-Cor Biosciences), respectively. The Li-Cor Odyssey® imaging system and software was used for antibody detection and quantification.
5
Protein Expression and Immunostaining Protocol
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All DNA constructs were transfected into 293T cells using Fugene 6 (Promega) and harvested 48 h later. An anti-Flag M2 affinity gel (Sigma, A2220) and an HA clone 16B12 monoclonal antibody (Covance, MMS-101R) were used for immunoprecipitation. The following primary antibodies were used for immunoblotting: mouse anti-TWIST1 2c1a (Santa Cruz Biotechnology), mouse anti-E-cadherin (BD, 610182), mouse anti-vimentin V9 (Thermo Scientific, MS-129-P), chicken anti-actin (Abcam, ab13822), anti-Flag (Sigma, F3165), anti-HA (Genetex, GTX115044), and anti-Myc hybridoma supernatant (clone 9E10).
Immunofluorescence was performed in eight-chamber slides by seeding 10,000 cells per well. Cells were fixed with 4% paraformaldehyde for 30 min and blocked in 5% goat serum/PBS-Tween for 1 h. Both primary and secondary antibodies were incubated in 5% goat serum/PBS-Tween overnight and for 1 h, respectively. The primary antibodies used were mouse anti-TWIST1 2c1a (Santa Cruz Biotechnology), rabbit anti-E-cadherin (Cell Signaling, 3195S), mouse anti-vimentin V9 (Thermo Scientific, MS-129-P), and nuclear DAPI staining (VectorShield). All images were imaged with an Olympus FV-1000 confocal microscope.
Immunofluorescence was performed in eight-chamber slides by seeding 10,000 cells per well. Cells were fixed with 4% paraformaldehyde for 30 min and blocked in 5% goat serum/PBS-Tween for 1 h. Both primary and secondary antibodies were incubated in 5% goat serum/PBS-Tween overnight and for 1 h, respectively. The primary antibodies used were mouse anti-TWIST1 2c1a (Santa Cruz Biotechnology), rabbit anti-E-cadherin (Cell Signaling, 3195S), mouse anti-vimentin V9 (Thermo Scientific, MS-129-P), and nuclear DAPI staining (VectorShield). All images were imaged with an Olympus FV-1000 confocal microscope.
6
Immunoblotting and Immunofluorescence Antibody Protocols
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Primary antibodies used for immunoblotting: anti-ACTB (MBL, #M177-3, 1:3000), anti-EGFR (MBL, #MI-12-1, 1:3000), anti-LC3 (MBL, #PM036, 1:3000), anti-VPS41 (SCBT, #sc-377118, 1:1000), anti-VPS8 (Proteintech, 15079-1-AP, 1:2000), anti-VPS39 (SCBT, sc-514762, 1:1000), anti-VPS3 (TRAP-1) (SCBT, #sc-13134, 1:1000), anti-VPS18 (Proteintech, #10901-1-AP, 1:2000), anti-VPS11 (SCBT, sc-515094, 1:2000), anti-VPS16 (Proteintech, # 17776-1-AP, 1:2000), anti-VPS33A (NovusBio, #NBP2-20872, 1:1000), anti-FLAG (Sigma-Aldrich, #F1804, 1:1000), anti-HA (Gene Tex, #GTX18181, 1:2000), anti-His (MBL, #PM032, 1:1000), anti-Myc (MBL, #M192-3, 1:10000), anti-V5 (MBL, #M215-3, 1:2000), anti-FLAG HRP-conjugated (FUJIFILM-Wako, # 019-22394, 1:10000), anti-Myc HRP-conjugated (MBL, #M192-7, 1:10000), anti-HA HRP-conjugated (MBL, #M180-7, 1:10000). Secondary antibodies used for immunoblotting: HRP-conjugated anti-rabbit IgG (Invitrogen, #31460, 1:3000 or 1:10000) and anti-mouse IgG (Invitrogen, #31430, 1:3000 or 1:10000).
Primary antibodies used for immunofluorescence staining: anti-LC3 (MBL, #PM036, 1:500. Secondary antibodies used for immunofluorescence staining: Alexa Fluor 488 anti-rabbit IgG (H + L) (Molecular Probes, #A21206).
Primary antibodies used for immunofluorescence staining: anti-LC3 (MBL, #PM036, 1:500. Secondary antibodies used for immunofluorescence staining: Alexa Fluor 488 anti-rabbit IgG (H + L) (Molecular Probes, #A21206).
7
Western Blot Antibody Panel for Protein Analysis
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The commercial antibodies used for Western blot analysis are as follows: anti-Twist1 (1:1000, sc-81417, Santa Cruz Biotechnology), anti-USP13 (1:3000, ab109264, Abcam), anti-RNF8 (1:1000, sc-271462, Santa Cruz Biotechnology), anti-FBXL14 (1:1000, HPA053889, Sigma-Aldrich), anti-βTrcp1 (1:1000, sc-390629, Santa Cruz Biotechnology), anti-E-cadherin (1:1000, sc-8426, Santa Cruz Biotechnology), anti-N-cadherin (1:1000, sc-59987, Santa Cruz Biotechnology), anti-Vimentin (1:1000, sc-6260, Santa Cruz Biotechnology), anti-MMP9 (1:1000, sc-21733, Santa Cruz Biotechnology), anti-Ubiquitin (1:1000, sc-8017, Santa Cruz Biotechnology), anti-Flag (1:3000, GTX115043, GeneTex), anti-GFP (1:3000, GTX113617, GeneTex), anti-HA (1:3000, GTX115044, GeneTex), anti-c-Myc (1:3000, PLA0001, Sigma), anti-GST (1:3000, sc-138, Santa Cruz Biotechnology), anti-GAPDH (1:3000, GTX627408, GeneTex), anti-tubulin (1:3000, bs-0715R, Bioss), and anti-Ubiquitin K48-specific antibody (1:1000, ZRB2150, Sigma-Aldrich). anti-Flag Affinity Gel (B23102), Poly Flag Peptide (B23111) and Protein A/G mix magnetic beads (B23202) were obtained from Bimake Chemicals. Cycloheximide (S7418) and MG132 (S2619) were obtained from Selleck Chemicals. TGF-β1 (100–21) was obtained from Peprotech.
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