Anti sigma1r

Manufactured by Merck Group

Anti-Sigma1R is a laboratory reagent used to study the sigma-1 receptor (Sigma1R), a protein involved in various cellular processes. The product is designed to selectively bind and inhibit the Sigma1R, allowing researchers to investigate its function and role in different biological systems.

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3 protocols using anti sigma1r

Subcellular Fractionation and Western Blot Analysis

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The isolated ER, c-mito, p-mito and MAM fractions were resuspended in lysis buffer, separated on SDS-polyacrylamide gel, and blotted onto PVDF membranes. Western blots were probed with antibodies against specific protein markers to verify the purity of the fractions and the presence of frataxin. The primary antibodies used were anti-ATP5A (Abcam, Cambridge, United Kingdom, ab110273), anti-Sigma1R (Sigma-Aldrich, St. Louis, Missouri, U.S., HPA018002), anti-GRP75 (Santa Cruz Biotechnology, Dallas, Texas, U.S., sc-13967), anti-VDAC1 (Abcam, Cambridge, United Kingdom, ab14734) and anti-FXN (Abcam, Cambridge, United Kingdom, ab110328). Secondary antibody (anti-mouse or anti-rabbit IgG) coupled to peroxidase was used for detection of the reaction with Amersham ECL Prime (GE Healthcare, Chicago, Illinois, U.S.). Imaging was performed with an ImageQuant LAS4000 (GE Healthcare, Chicago, Illinois, U.S.).

Rodríguez L.R., Calap-Quintana P., Lapeña-Luzón T., Pallardó F.V., Schneuwly S., Navarro J.A, & Gonzalez-Cabo P. (2020). Oxidative stress modulates rearrangement of endoplasmic reticulum-mitochondria contacts and calcium dysregulation in a Friedreich's ataxia model. Redox Biology, 37, 101762.

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Plasmid Construction and Antibody Details

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Plasmids expressing cMyc-tagged mouse SEPN1 (with A1284T/C conversion of the TGA selenocysteine codon into Cys to increase expression) and cMyc-tagged human Desmin were constructed in a CMV promoter-derived mammalian expression vector.
Expression plasmids encoding N-terminally FLAG-tagged human SEPN1 and the mutants SEPN1 SS (C427S, U428S) and SEPN1 CC (U428C) were constructed in pSel-Express vector (a kind gift from Vladimir Gladyshev) [16 ].
The primary antibodies used were: anti-VDAC2, anti-Cyt C, anti-Calnexin, anti-SERCA2, anti-RyR, anti-cMyc, and anti-PDI from Abcam; anti-Flag, anti-β-tubulin, and anti-Sigma 1-R from Sigma-Aldrich; anti-IP3R3 and Cyt c from BD Biosciences; anti-Tom40 from Santa Cruz, and homemade rabbit anti-SEPN1 [21 (link)]. Secondary antibodies Alexa Fluor 488 and 594 were from Life Technologies.

Filipe A., Chernorudskiy A., Arbogast S., Varone E., Villar-Quiles R.N., Pozzer D., Moulin M., Fumagalli S., Cabet E., Dudhal S., De Simoni M.G., Denis R., Vadrot N., Dill C., Giovarelli M., Szweda L., De Palma C., Pinton P., Giorgi C., Viscomi C., Clementi E., Missiroli S., Boncompagni S., Zito E, & Ferreiro A. (2020). Defective endoplasmic reticulum-mitochondria contacts and bioenergetics in SEPN1-related myopathy. Cell Death and Differentiation, 28(1), 123-138.

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Subcellular Fraction Protein Analysis

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The isolated ER, c-mito, p-mito and MAM fractions were resuspended in lysis buffer, separated on SDS-polyacrylamide gel, and blotted onto PVDF membranes. Western blots were probed with antibodies against specific protein markers to verify the purity of the fractions and the presence of frataxin. The primary antibodies used were anti-ATP5A (Abcam, ab110273), anti-Sigma1R (Sigma, HPA018002), anti-GRP75 (Sta Cruz Biotec, sc-13967), anti-VDAC1 (Abcam, ab14734) and anti-FXN (Abcam, ab110328). Secondary antibody (anti-mouse or anti-rabbit IgG) coupled to peroxidase was used for detection of the reaction with Amersham ECL Prime. Imaging was performed with an ImageQuant LAS4000 (GE Healthcare).

Rodríguez L.R., Calap-Quintana P., Lapeña-Luzón T., Pallardó F.V., Schneuwly S., Navarro J.A., & González‐Cabo P. (2020). Disarrangement of Endoplasmic reticulum-mitochondria communication impairs Ca²⁺ homeostasis in FRDA.

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