Asc 013

Manufactured by Alomone

ASC-013 is a chemical compound sold as a research tool. It is used in laboratory settings for in vitro and in vivo studies. The core function of ASC-013 is to serve as a research tool to investigate cellular processes and signaling pathways.

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Lab products found in correlation

Pierce cell surface protein isolation kit, by Thermo Fisher Scientific (2 mentions) Pierce ecl immunoblotting substrate, by Thermo Fisher Scientific (2 mentions) Halt protease inhibitor cocktail, by Thermo Fisher Scientific (2 mentions) Asc 005, by Alomone (2 mentions) Apr 003, by Alomone (2 mentions) Protein g sepharose fast flow, by Cytiva (1 mentions) Hrp conjugated goat anti rabbit secondary antibody, by Promega (1 mentions)

3 protocols using asc 013

Cardiac Protein Interactions via Immunoprecipitation

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LV myocardium was mechanically homogenized (using a stainless-steel
pestle) in Tris buffer containing (in mmol/L): 50 Tris-HCl, 200 NaCl, 20
NaF, 1 Na₃VO₄, 1 DTT [pH 7.4] and protease inhibitor
cocktail. Protein concentration was determined by BCA assay. The homogenate
(1 mg protein) was suspended in dilution medium containing (in mmol/L): 50
Tris-HCl, 154 NaCl, 1% CHAPS, 1 NaF, 1 Na₃VO₄ [pH 7.4]
and protease inhibitor cocktail. CASK, Na_V1.5 or CaMKII,
respectively, were immunoprecipitated with rabbit polyclonal anti-CASK (1.4
mg/ml, Abcam, discontinued, AB11343), rabbit polyclonal
anti-Na_V1.5 (1 mg/ml, Alomone, ASC-013) or rabbit polyclonal
anti-CaMKII antibody (1 mg/ml^{32 (link)}, Don Bers' lab) by preincubation at 4°C
overnight in protein G–sepharose Fast Flow (prewashed; 2 hours,
4°C; Amersham Biosciences). As control, rabbit polyclonal
anti-calsequestrin (1 mg/ml, Thermo Scientific) was used. After
centrifugation, pellets were washed with Tris buffer containing (in mmol/L):
50 Tris-HCl, 154 NaCl [pH 7.4], and immunoprecipitated proteins were eluted
in 2X Laemmli sample buffer containing 4% β-mercaptoethanol (30
minutes, 37°C) followed by centrifugation. Supernatants were
subjected to Western blotting.

Mustroph J., Sag C.M., Bähr F., Schmidtmann A.L., Gupta S.N., Dietz A., Towhidul Islam M.M., Lücht C., Beuthner B.E., Pabel S., Baier M.J., El-Armouche A., Sossalla S., Anderson M.E., Moellmann J., Lehrke M., Marx N., Mohler P.J., Bers D.M., Unsöld B., He T., Dewenter M., Backs J., Maier L.S, & Wagner S. (2021). Loss of CASK Accelerates Heart Failure Development. Circulation research.

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Western Blot Analysis of Ion Channels

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Oocytes expressing full-length receptors or different combinations of the split-intein receptor fragment fusion proteins were isolated 3–4 days after RNA injection and washed twice with phosphate-buffered saline (PBS). Total cell lysates were obtained by lysing the oocytes in Pierce™ IP lysis buffer with added Halt protease inhibitor cocktail (Thermo Fisher Scientific). Surface proteins were purified with the Pierce™ Cell Surface Protein Isolation Kit (Thermo Fisher Scientific). Purified surface proteins or total cell lysates were run on a 4–12% BIS-TRIS gel (for P2X2) or 3–8% Tris-acetate gel (for Na_V1.5) and transferred to a polyvinylidene difluoride membrane. Membranes were incubated with rabbit polyclonal anti-Na_V1.5 (#ASC-005, Alomone labs; 1:2000), anti-Na_V1.5 (#ASC-013, Alomone labs; 1:1500) or anti-P2X2 Antibody (#APR-003, Alomone labs; 1:2000) and the bound primary antibodies were detected by a HRP-conjugated goat anti-rabbit secondary antibody (W401B, Promega; 1:2000). Membranes were developed and visualized using the Pierce™ ECL immunoblotting substrate (Thermo Fisher Scientific).

Khoo K.K., Galleano I., Gasparri F., Wieneke R., Harms H., Poulsen M.H., Chua H.C., Wulf M., Tampé R, & Pless S.A. (2020). Chemical modification of proteins by insertion of synthetic peptides using tandem protein trans-splicing. Nature Communications, 11, 2284.

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Receptor Expression and Detection

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Oocytes expressing full-length receptors or different combinations of the split-intein receptor fragment fusion proteins were isolated 3-4 days after RNA injection and washed twice with PBS. Total cell lysates were obtained by lysing the oocytes in Pierce™ IP lysis buffer with added Halt protease inhibitor cocktail (Thermo Fisher Scientific). Surface proteins were purified with the Pierce™ Cell Surface Protein Isolation Kit (Thermo Fisher Scientific).
Purified surface proteins or total cell lysates were run on a 4-12 % BIS-TRIS gel (for P2X2) or 3-8 % Tris-acetate gel (for NaV1.5) and transferred to a PVDF membrane. Membranes were incubated with rabbit polyclonal anti-NaV1.5 (#ASC-005, Alomone labs; 1:2000), anti-NaV1.5 (#ASC-013, Alomone labs; 1:1500) or anti-P2X2 Antibody (#APR-003, Alomone labs; 1:2000) and the bound primary antibodies were detected by a HRP-conjugated goat anti-rabbit secondary antibody (W401B, Promega; 1:2000). Membranes were developed and visualized using the Pierce™ ECL immunoblotting substrate (ThermoFisher Scientific).

Khoo K.K., Galleano I., Gasparri F., Wieneke R., Harms H.J., Poulsen M.H., Chua H.C., Wulf M., Tampé R., & Pless S.A. (2020). Chemical modification of proteins by insertion of synthetic peptides using tandem protein trans-splicing.

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