Anti human actin

Manufactured by Merck Group

Sourced in United States

Anti-human actin is a laboratory reagent used to detect and quantify the presence of actin, a ubiquitous cytoskeletal protein, in human samples. It is a highly specific antibody that binds to actin, allowing researchers to study the distribution and dynamics of this important structural component in cells.

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4 protocols using anti human actin

Immunodetection of Innate Immune Receptors

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Antibodies anti-Human CD81 mouse mAb (clone 1.3.3.22, Santa Cruz Biotechnology, Santa Cruz CA); Anti-human CD4 mouse mAb (clone SK3) and an isotype control (clone MOPC-21, Biolegend, San Diego CA); Anti-human TLR3 rabbit mAb (clone D10F10, Cell Signaling Technology, Danvers MA); Anti-human TLR7 rabbit pAb (#2633S, Cell signaling Technology); Anti-human TLR8 rabbit mAb (clone D3Z6J, Cell Signaling Technology); Anti-human TLR9 rabbit mAb (clone D2C9, Cell Signaling Technology); Anti-human MyD88 rabbit mAb (clone D80F5, Cell Signaling Technology); Anti-human TRIF/TICAM-1 (clone MAB6216, R&D Systems, Minneapolis MN) mouse mAb; Anti-human NALP3 rabbit pAb (Imgenex, San Diego CA); Anti-human RIG-I (D14G6) rabbit mAb (clone D14G6, Cell Signaling Technology); Anti-human AIM2 rabbit pAb (Abcam, Cambridge, MA) and Anti-human Actin (A2066, Sigma-Aldrich) were purchased from respective vendors. Secondary Antibodies anti-mouse IgG-HRP goat polyclonal (HAF007, R&D Systems), anti-rabbit IgG HRP-linked (7074, Cell Signaling Technology) were also purchased.

Chattergoon M.A., Latanich R., Quinn J., Winter M.E., Buckheit RW 3.r.d., Blankson J.N., Pardoll D, & Cox A.L. (2014). HIV and HCV Activate the Inflammasome in Monocytes and Macrophages via Endosomal Toll-Like Receptors without Induction of Type 1 Interferon. PLoS Pathogens, 10(5), e1004082.

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Erythrocyte Ghost Protein Analysis

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Erythrocyte ghosts were isolated from fresh human blood via hypotonic lysis of erythrocytes in 20 mM sodium phosphate (pH 7.4) and supplemented with 0.1% saponin, as described previously [24 (link)]. The purified ghosts were incubated with 50 nM of each enzyme at pH 5.0 or 7.0 at 37 °C for 4 h. The mixtures were analyzed by SDS–PAGE, followed by Coomassie blue staining. For immunoblot analysis, the mixtures were transferred to nitrocellulose membranes (0.45 µm: Bio-Rad, Hercules, CA, USA), and the membranes were blocked with phosphate-buffered saline (PBS, pH 7.4), supplemented with 0.05% Tween 20 and 5% (w/v) skim milk for 1 h at room temperature. The membranes were rinsed with PBS, supplemented with 0.05% Tween 20 (PBST) several times, and incubated with one of the monoclonal antibodies against anti-human spectrin (Sigma, 1:500 dilution), anti-human actin (Sigma, 1:1000 dilution), and anti-human band 3 (Sigma, 1: 5000 dilution) for 3 h at room temperature, respectively. Each membrane was repeatedly washed with PBST and incubated with horseradish peroxidase-conjugated host-specific antibodies (Sigma) for 2 h at room temperature. The immunoreactive bands were visualized with the SuperSignal West Pico PLUS chemiluminescent substrate (Thermo Fisher Scientific).

Lê H.G., Kang J.M., Võ T.C., Yoo W.G., Lee K.H, & Na B.K. (2022). Biochemical Properties of Two Plasmodium malariae Cysteine Proteases, Malapain-2 and Malapain-4. Microorganisms, 10(1), 193.

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Monoclonal Antibody and Kinase Inhibitor Assay

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Anti-human-IDO monoclonal antibody was prepared and utilized as previously reported (8 (link)). Anti-human-actin(Sigma), anti-mouse-CD49b(R&DSystems), anti-HGF-α (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA), anti-phospho-c-Met, anti-c-Met, anti-phospho-AKT, anti-AKT, anti-phospho-ERK, anti-ERK, anti-phospho-STAT3 and anti-STAT3 (Cell Signaling Technology, Inc., Danvers, MA, USA) antibodies were purchased and utilized according to the manufacturer's instructions.
The c-Met tyrosine kinase inhibitor PHA-665752((3Z)-5-[(2,6-dichlorobenzyl)sulfonyl]-3-[(3,5-dimethyl-4-{[(2R)-2-(pyrrolidin-1-ylmethyl)pyrrolidin-1-yl]carbonyl}-1H-pyrrol-2-yl)methylene]-1,3-dihydro-2H-indol-2-one; Merck KGaA, Darmstadt, Germany) (28 (link)), the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 (2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; Cell Signaling Technology) (29 (link)), the MEK1/2 inhibitor U0126 (1,4-diamino-2,3-dicyano-1,4-bis[2-aminophenylthio] butadiene; Cell Signaling Technology) (30 (link)), and the STAT3 inhibitor WP1066 ((2E)-3-(6-Bromo-2-pyridinyl)-2-cyano-N-[(1S)-1-phenylethy]-2-propenamide P; Santa Cruz Biotechnology) (31 (link)) were purchased and were utilized according to the corresponding manufacturer's instructions.

WANG D., SAGA Y., SATO N., NAKAMURA T., TAKIKAWA O., MIZUKAMI H., MATSUBARA S, & FUJIWARA H. (2016). The hepatocyte growth factor antagonist NK4 inhibits indoleamine-2,3-dioxygenase expression via the c-Met-phosphatidylinositol 3-kinase-AKT signaling pathway. International Journal of Oncology, 48(6), 2303-2309.

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Tyrosine Hydroxylase Protein Detection

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Western blots were performed as described earlier {Malard, 2009 #524}. After 7 days of differentiation, the cells were trypsinized, washed with PBS, then incubated with RIPA buffer (Santa Cruz) containing protease inhibitor cocktail tablet (Complete EDTA free TM Roche) and phosphatase inhibitor (Cocktail I and II, Sigma) . The cellular extracts were lysed using a Dounce homogenizer (20 strokes), incubated on ice for 60 min, and then centrifuged at 16,000 g for 60 min. The protein content of the supernatants was determined with the Coomassie Protein Assay (Thermo Scientifc). Proteins were separated on a 4-12% NuPAGE® Bis-Tris gel (Invitrogen) in a MOPS/SDS Running Buffer (Invitrogen) at 200 V. Proteins from the gel were electroblotted on a 0.45 µm PVDF membrane (Thermo scientific) using a semi-dry electroblot system (Biometra). After a saturation step in milk 5%, the membrane was incubated overnight at 4°C in a 5 % milk solution containing a polyclonal Antibody Rabbit anti-Tyrosine Hydroxylase (1/1000, cat# AB152, Millipore). Primary antibody was then detected using an anti Rabbit HRP (1/10000, Sigma; A6154). A mouse monoclonal anti-human actin from Sigma ref A3854 was used as a control at 1/25000. The signals were revealed using the Western Chemiluminescent HRP Substrate from Millipore.

Carmona A., Malard V., Avazeri E., Roudeau S., Porcaro F., Paredes E., Vidaud C., Bresson C, & Ortega R. (2018). Uranium exposure of human dopaminergic cells results in low cytotoxicity, accumulation within sub-cytoplasmic regions, and down regulation of MAO-B. Neurotoxicology, 68.

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