Mouse anti ha

Manufactured by Roche

Sourced in United States, Switzerland

The Mouse anti-HA is a monoclonal antibody that specifically recognizes the hemagglutinin (HA) epitope tag. It is widely used in research applications to detect and purify HA-tagged proteins.

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Lab products found in correlation

Mouse anti flag m2, by Merck Group (4 mentions) Mouse anti v5, by Thermo Fisher Scientific (3 mentions) Mouse anti ha, by BioLegend (3 mentions) Prolong gold antifade reagent with dapi, by Thermo Fisher Scientific (3 mentions) Rabbit anti tapin1, by Roche (2 mentions) Phallo din tritc, by Thermo Fisher Scientific (2 mentions) Inverted 6000, by Leica (2 mentions) Fibronectin coated slides, by Merck Group (2 mentions) Mouse anti flag, by Merck Group (2 mentions) Rabbit anti flag, by Merck Group (2 mentions) Rabbit anti gfp, by Abcam (2 mentions) Goat anti mouse alexa 488, by Thermo Fisher Scientific (2 mentions) Irdye 680lt goat anti rabbit, by LI COR (2 mentions) Irdye 800cw goat anti mouse, by LI COR (2 mentions) Alexa fluor 488, by Thermo Fisher Scientific (2 mentions) Mini protean tgx precast protein gel, by Bio-Rad (1 mentions) Rabbit anti akt, by Cell Signaling Technology (1 mentions) Rabbit anti erk1 2, by Cell Signaling Technology (1 mentions) Peroxidase conjugated sheep anti mouse secondary antibodies, by Jackson ImmunoResearch (1 mentions) Chicken anti gfp, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Mouse anti-HA clone 12CA5 (6 citations) Anti HA mouse primary antibody (2 citations) Mouse anti-HA (12CA5, (15 citations) Mouse anti-HA antibody 12CA5 (2 citations) Mouse monoclonal anti-HA antibody (12CA5, (5 citations) Mouse monoclonal anti-HA (4 citations) Mouse monoclonal anti-HA antibody (11 citations) Mouse anti-HA antibody (7 citations) Mouse anti-HA tag (5 citations) Mouse anti-HA 12CA5 monoclonal antibodies (2 citations)

35 protocols using mouse anti ha

Detecting Protein-Protein Interactions via IP

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For detecting interaction between Pes4–3HA and Bmh1-GFP, the immunoprecipitation protocol described for Bmh1-GFP was used with the following modifications: 1) 4μg anti-HA mouse (12CA5 Roche, Catalog # 11583816001) used for immunoprecipitating Pes4–3HA. 2) For inputs, 25μL protein sample was mixed with 5μL 3X SDS buffer and boiled for 5 mins. 25μL input sample was loaded in the gel.
For detecting interaction between Pes4–3HA and LacI-GFP, the immunoprecipitation protocol described for Pes4–3HA was used with the following modifications: 50μM copper sulfate was added in SPM at 0 hour for expressing LacI-GFP. After loading the samples in 4–20% Mini-PROTEAN TGX Precast protein gel (BIO-RAD, Catalog #4561094), the samples were resolved at 150V for 90 minutes and transferred onto a PVDF membrane.
The primary antibodies used were rabbit anti-GFP (generous gift of Dr. Claire Walczak, 1:10000), and anti-HA mouse (12CA5 Roche, Catalog # 11583816001, 1:1000). The secondary antibodies used were ECL-anti-rabbit HRP IgG (GE Healthcare, NA9340V, 1:10000), and ECL-anti-mouse HRP IgG (GE Healthcare, NA9310V, 1:10000).

Gavade J.N., Puccia C.M., Herod S.G., Trinidad J.C., Berchowitz L.E, & Lacefield S. (2022). Identification of 14-3-3 Proteins, Polo kinase, and RNA-binding protein Pes4 as Key Regulators of Meiotic Commitment in Budding Yeast. Current biology : CB, 32(7), 1534-1547.e9.

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Protoplast Transfection and Immunodetection

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After an overnight incubation, the ten 30 µL protoplast transfection reactions were pooled, diluted 10-fold v/v with W5 (154 mM NaCl; 125 mM CaCl₂; 5 mM KCl; 5 mM glucose) and pelleted at 50 g for 20 min, 4 °C. The cells were moved to a 2 mL Eppendorf, and repelleted at 100 g for 10 min, 4 °C, to remove any residual buffer. Approximately 100 µL of glass beads (∅ 0.1 mm) and 125 µL of Lyse and Load Buffer (50 mM Tris/HCl pH 6.8; 4% SDS; 8M Urea; 30% v/v glycerol; 0.1 M DTT; 0.005% Bromophenol blue) were added, vigorously vortexed for 2 min, incubated at 65 °C for 10 min, then centrifuged for 10 min 4 °C at 8000× g. The supernatant used for SDS-PAGE and wet immunoblotting onto PVDF. The HA epitope was detected with anti-HA mouse (Roche) antibody followed by an AP-conjugated anti-mouse (BioRad).

Rieger J., Fitz M., Fischer S.M., Wallmeroth N., Flores-Romero H., Fischer N.M., Brand L.H., García-Sáez A.J., Berendzen K.W, & Mira-Rodado V. (2023). Exploring the Binding Affinity of the ARR2 GARP DNA Binding Domain via Comparative Methods. Genes, 14(8), 1638.

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Western Blot Analysis of HEK293 Cells

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HEK293 cells were lysed for 1 hour at 4° C in TNE lysis buffer containing (in mM): 50 Tris/HCl (pH 8.0), 150 NaCl, 5 EDTA, 1% (v/v) Triton X-100 and protease inhibitors (pepstatin 1 μg/ml, PMSF 1 mM, leupeptin 5 μg/ml and aproptin 5 μg/ml). Protein lysates were denatured in Laemmli containing 100 mM dithiothreitol (DTT, 30 minutes, 37°C) and subsequently subjected to SDS-PAGE. Immunoblots were incubated with mouse anti-HA (Roche, high affinity 3F10, 1:5,000), rabbit anti-Akt (Cell signaling, 1:1000) and rabbit anti-ERK1/2 (Cell signaling, 1:1,000) primary antibodies and peroxidase conjugated sheep anti-mouse secondary antibodies (Jackson Immunoresearch, 1:10,000).

Blanchard M.G., de Baaij J.H., Verkaart S.A., Lameris A.L., Basmadjian C., Zhao Q., Désaubry L., Bindels R.J, & Hoenderop J.G. (2015). Flavaglines Stimulate Transient Receptor Potential Melastatin Type 6 (TRPM6) Channel Activity. PLoS ONE, 10(3), e0119028.

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Antibody Toolkit for Neuroscience Research

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The following primary antibodies were used: rat, anti-HA (Roche); rabbit, anti-Myc (Cell Signaling Technologies); rabbit anti-DsRed (Takara 632496); rabbit anti-Synapsin 1 (Invitrogen A6442); rabbit anti-Neuroligin-2 (Synaptic Systems 129203) rabbit anti-VGAT (Chemicon AB5855); mouse anti-GFP (Molecular Probes A11120) mouse anti-Gephyrin (Synaptic Systems 147 011); mouse anti-HA (Roche 11 583 816); rabbit anti-synapsin-1XP (Cell Signaling); guinea pig anti-VGAT (Chemicon); Chicken anti-GFP (Invitrogen A10262). Animal-specific fluorescently-tagged secondary antibodies from Invitrogen were used. HRP-tagged secondaries for western blots: anti-mouse (Pierce 31430), anti-rabbit (Jackson ImmunoResearch 111-035-003). The mouse anti-Myc-9E10 monoclonal antibody was obtained from the Developmental Studies Hybridoma Bank, created by the NICHD of the NIH and maintained at The University of Iowa, Department of Biology, Iowa City, IA 52242.

Steffen D.M., Ferri S.L., Marcucci C.G., Blocklinger K.L., Molumby M.J., Abel T, & Weiner J.A. (2021). The γ-Protocadherins interact physically and functionally with Neuroligin-2 to negatively regulate inhibitory synapse density and are required for normal social interaction. Molecular neurobiology, 58(6), 2574-2589.

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Antibody Sources and Immunostaining Protocols

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Antibodies were from the following sources: rat anti-Yl and rabbit anti-Yolk antibodies were from Dr. Mahowald [48 (link)]. mouse anti-Rab7 (1:10, Developmental Studies Hybridoma Bank (DSHB)), mouse anti-Rab11 (1:200, BD Transduction Laboratories (#610657)), mouse anti-actin (1:10000, MAB1501, Chemicon, and #ab-6276, Abcam); mouse anti-αTubulin (1:10000, DM1A, Sigma); mouse anti-HA (12CA5, Roche); chicken anti-GFP (Aves. 1:10,000 for Western blot and 1:2,000 for immunofluorescent staining). Alexa Fluor 488 AffiniPure Donkey Anti-Chicken IgY (1:1000, 703-545-155) and Rhodamine Red AffiniPure Donkey Anti-Mouse IgG (1:1000, 715-295-151) from Jackson ImmunoResearch Inc. TRITC-Phalloidin (Phalloidin–Tetramethylrhodamine B isothiocyanate) (1:200, P1951, Sigma). Alexa Fluor 488-Phalloidin (1:200, A-12379), Alexa Fluor 594-Phalloidin (1:200, A-12381), Alexa Fluor 647-Phalloidin (1:200, A-30107), Alexa Fluor 680-(A-21076) were from Molecular Probes (Invitrogen). Alexa Fluor 680-(A-21076) and Alexa Fluor 800-(926–32212) conjugated secondary antibodies for immunoblotting (1:10,000) were from Molecular Probes (Invitrogen) and LI-COR, respectively.

Yu Y., Chen D., Farmer S.M., Xu S., Rios B., Solbach A., Ye X., Ye L, & Zhang S. (2024). Endolysosomal trafficking controls yolk granule biogenesis in vitellogenic Drosophila oocytes. PLOS Genetics, 20(2), e1011152.

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Immunofluorescence Staining of SN56 Cells

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Antibodies were: Mouse Anti-HA (Roche Applied Science), Rabbit Anti-Arf6 (Sigma), Rabbit Anti-Arf1 (Epitomics), Rabbit Anti-Rac1 (Santa Cruz Biotechnology). SN56 cells were obtained from Dr. Jane Rylett. Fluorescently-labeled secondary antibodies and Zenon® Alexa Fluor® 647 Mouse IgG2b Labeling Kit were purchased from Life Technologies (California) Fluorescent dextran (Cascade blue, 3000 Da, lysine fixable) was purchased from Life Technologies (California). ELISA assays were from Life Technologies (California). Dulbecco’s modified Eagle’s medium, fetal bovine serum, Hank’s balanced salt solution (HBSS), penicillin, streptomycin, trypsin and neurobasal media were all purchased from Gibco.

Tang W., Tam J.H., Seah C., Chiu J., Tyrer A., Cregan S.P., Meakin S.O, & Pasternak S.H. (2015). Arf6 controls beta-amyloid production by regulating macropinocytosis of the Amyloid Precursor Protein to lysosomes. Molecular Brain, 8, 41.

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Immunofluorescence Analysis of Cellular Proteins

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BL3 and TBL3 cells were plated on Fibronectin coated slides (Sigma; Ref: F1141). NIH/3T3 cells and Mouse Immortalized Fibroblasts cells-transfected by indicated constructs were plated on slides. All cells were then fixed in PBS 3.7% Formaldehyde for 15min at room temperature. Slides with bovines cells or NIH/3T3 cells were rinsed in PBS and permeabilized with PBS 0.2% Triton X-100 for 5min and then blocked for 30min with PBS 1% SVF and 1% BSA to prevent non-specific staining. These slides were incubated with rabbit anti-TaPin1 (1/250) and/or mouse anti-HA (1/1000, Roche, Ref: 11583816001) in PBS 1% SVF and 1% BSA at room temperature for 40min. After washing in PBS 0.2% Tween, the slides were incubated with Texas Red dye-conjugated AffinyPure Donkey anti-rabbit IgG and/or Cy2 AffinyPure Donkey anti-mouse IgG (1/5000, Jackson Immunology, Ref: 711-075-152 or Ref: 715-225-150) for 30min. Slides with Murine Immortalized Fibroblasts cells were incubated for 15min with Phalloïdin-TRITC (Life Technologies, Ref: R415). All slides were subsequently washed in PBS 0.2% Tween, mounted on slides and covered with ProLong Gold Antifade Reagent with DAPI (Invitrogen, Ref: P-36931). Images of immunofluorescence staining (Mouse Immortalized Fibroblasts cells) were photographed with a fluorescent microscope (Leica Inverted 6000). Staining was repeated for three independent biological replicates.

Marsolier J., Perichon M., DeBarry J., Villoutreix B., Chluba J., Lopez T., Garrido C., Zhou X., Lu K., Fritsch L., Ait-Si-Ali S., Mhadhbi M., Medjkane S, & Weitzman J. (2015). Theileria parasites secrete a prolyl isomerase to maintain host leukocyte transformation. Nature, 520(7547), 378-382.

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Immunofluorescence Analysis of Cellular Proteins

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Protein Fractionation and Western Blot Analysis

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Protein fractionation was performed with modifications according to Till et al (49) and
Baumgarten et al (12) . Synchronous middle-late stage parasites were released from 1 mL infected red blood cells by 0.15% saponin lysis and washed twice in 1 × pre-cooled PBS.
The parasite pellet was first resuspended in 0.5 mL cytoplasmic lysis buffer (20mM Hepes pH7.9, 10mM KCl, 1.5mM MgCl2, 1mM EDTA, 1mM EGTA, 0.65% NP-40, 1mM DTT, 1x Protease inhibitor cocktail) and incubated for 30 min on ice. Parasites were further homogenized for 100 strokes in a glass douncer, and the supernatant containing cytoplasmic proteins was isolated via centrifugation (10,000 rpm, 10 min, 4 °C). Protein samples were separated on SDS-PAGE and transferred onto PVDF membrane and visualized using ECL Western Blotting Detection Kit (GE Healthcare). The primary antibodies used in this study were mouse anti-HA (1:2000, Roche) and anti-GAPDH
(1:1000, Servicebio, cat# GB12002), and secondary antibody was HRP-conjugated goat anti-mouse IgG (1:5000, Abcam, cat# ab6789) .

Liu M., Guo G., Qian P., Mu J., Lu B., He X., Shang X., Yang G., Shen S., Liu W., Wang L., Gu L., Mu Q., Yu X., Zhao Y., Culleton R., Cao J., Jiang L., Wellems T.E., Yuan J., Jiang C., & Zhang Q. (2021). 5-methylcytosine modification byPlasmodiumNSUN2 stabilizes mRNA and mediates the development of gametocytes.

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Isolation and Western Blot Analysis of Fly Mitochondria

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The isolation of fly mitochondria was performed as previously described,^{56 (link)} except that 20 eyes were gently homogenized in 1 ml of mitochondrial isolation buffer (250 mM sucrose, 10 mM Tris (pH 7.4), and 0.15 mM MgCl₂). The mitochondrial (pellet) and cytosolic (supernatant) fractions were solubilized in a 50 μl SDS sample buffer and then subjected to western blotting.
In order to detect HA-tagged PINK1 and Flag-tagged Parkin on western blots, the samples were homogenized in an SDS sample buffer, fractionated by SDS-PAGE, and transferred to Immobilon-FL membranes (Millipore, Billerica, MA, USA). The blots were then incubated with mouse anti-HA (Roche, Indianapolis, IN, USA) or mouse anti-Flag (Sigma) primary antibodies or control antibodies, which included mouse anti-Rh1 (Developmental Studies Hybridoma Bank, Ames, IA, USA), rabbit anti-CoIV (Abcam, Cambridge, MA, USA), and rat-anti-TOM20. The blots were subsequently incubated with IRDye 800-labeled anti-mouse IgG and IRDye 680-labeled anti-rabbit or rat IgG (LI-COR Biosciences, Lincoln, NE, USA). The signals were then detected using an Odyssey Infrared Imaging System (LI-COR Biosciences).

Huang Z., Ren S., Jiang Y, & Wang T. (2016). PINK1 and Parkin cooperatively protect neurons against constitutively active TRP channel-induced retinal degeneration in Drosophila. Cell Death & Disease, 7(4), e2179-.

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