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Ab2827

Manufactured by Abcam
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Ab2827 is a secondary antibody product from Abcam. It is a polyclonal antibody conjugated with a fluorescent dye. The core function of this product is to detect and visualize target proteins in various experimental applications.

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Lab products found in correlation

Ab2864, by Abcam (2 mentions) Ma3 919, by Thermo Fisher Scientific (2 mentions) Triton x 100, by Merck Group (1 mentions) A1 laser scanning confocal microscope, by Nikon (1 mentions) Anti grp78, by Cell Signaling Technology (1 mentions) Ma3 927, by Thermo Fisher Scientific (1 mentions) Anti gapdh, by Cell Signaling Technology (1 mentions) Anti α tubulin, by Cell Signaling Technology (1 mentions) Anti grp94, by Cell Signaling Technology (1 mentions) A 21245, by Thermo Fisher Scientific (1 mentions) 4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions) A11001, by Thermo Fisher Scientific (1 mentions) Dulbecco s phosphate buffered saline pbs, by Merck Group (1 mentions) Ab2861, by Abcam (1 mentions) Ma3 926, by Thermo Fisher Scientific (1 mentions) A11011, by Thermo Fisher Scientific (1 mentions) F1804, by Merck Group (1 mentions) A7811, by Merck Group (1 mentions) Ab2865, by Abcam (1 mentions) Ab176333, by Abcam (1 mentions)

6 protocols using ab2827

1

Immunofluorescence Staining of CD38 and RyR2

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Cells were fixed in in 4% paraformaldehyde/PBS for 15 min, washed in PBS (three changes, 5 min each), permeabilized using 0.1% Triton X-100 (Sigma-Aldrich) for 10 min, washed in PBS, and blocked with 1% bovine serum albumin for 60 min before being incubated with the primary antibody at 4 °C overnight. The next day, cells were first washed with PBS before being incubated with secondary antibody at room temperature for 2 h (either Alexa Fluor 488–conjugated donkey anti-rabbit or Dylight 550 donkey anti-mouse, 1:1000 dilution) and then washed. Finally, coverslips were mounted using Vectashield® and permanently sealed. Cells were stored in the dark at 4 °C and visualized within 2 days. For control experiments, we performed the same procedure with omission of the primary antibodies or with cells from CD38−/− mice. Primary antibodies against CD38 (sc-15362, Santa Cruz Biotechnology) and RyR2 (ab2827, Abcam) were used at 1:100 and 1:200 dilution, respectively. Observations were carried out using a Nikon A1 confocal laser-scanning microscope equipped with a ×60 objective, and images were processed using ImageJ software.
Lin W.K., Bolton E.L., Cortopassi W.A., Wang Y., O'Brien F., Maciejewska M., Jacobson M.P., Garnham C., Ruas M., Parrington J., Lei M., Sitsapesan R., Galione A, & Terrar D.A. (2017). Synthesis of the Ca2+-mobilizing messengers NAADP and cADPR by intracellular CD38 enzyme in the mouse heart: Role in β-adrenoceptor signaling. The Journal of Biological Chemistry, 292(32), 13243-13257.
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2

Antibody Validation for Cellular Imaging

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Primary rabbit polyclonal anti-REEP5 antibody (IB: 1:1000 dilution, IF: 1:800 dilution; 14643-1-AP; Proteintech), polyclonal anti-Nogo-A/B (IB: 1:1000 dilution, IF: 1:1000 dilution; PA1-41220; ThermoFisher), polyclonal anti-ATL3 antibody (IB: 1:1000 dilution, IF: 1:800 dilution; PA5-24652; ThermoFisher), polyclonal anti-CKAP4 antibody (IB: 1:1000 dilution, IF: 1:800 dilution; PA5-42926; ThermoFisher) and mouse monoclonal anti-Alpha sarcomeric actinin antibody (IF: 1:500 dilution; MA1-22863; ThermoFisher), monoclonal anti-Triadin antibody (IF: 1:500 dilution; MA3-927; ThermoFisher), monoclonal anti-RyR2 antibody (IF: 1:500 dilution; ab2827; Abcam) were used for immunocytochemistry and immunoblot studies. Anti-alpha tubulin (#2144), anti-GRp78 (#3177), anti-GRp94 (#2104), anti-GAPDH (#2118) antibodies from Cell Signaling; anti-ATF4 (ab216839), anti-Caspase12 (ab62484) antibodies from Abcam were used for immunoblot studies at 1:1000 dilution. Anti-GFP (sc390394) antibody from Santa Cruz Biotechnology was used for immunoblot studies at 1:1000 dilution.
Lee S.H., Hadipour-Lakmehsari S., Murthy H.R., Gibb N., Miyake T., Teng A.C., Cosme J., Yu J.C., Moon M., Lim S., Wong V., Liu P., Billia F., Fernandez-Gonzalez R., Stagljar I., Sharma P., Kislinger T., Scott I.C, & Gramolini A.O. (2020). REEP5 depletion causes sarco-endoplasmic reticulum vacuolization and cardiac functional defects. Nature Communications, 11, 965.
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3

Immunolabeling of Calcium Signaling Proteins

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Cells cultured on glass coverslips were washed once with Dulbecco's phosphate buffered saline (PBS, Sigma-Aldrich, MO, USA), fixed with 4% paraformaldehyde (in PBS) for 5 min and permeabilized with 0.5% Triton-X (in PBS) (Sigma-Aldrich, MO, USA) for 10 min. Coverslips were washed twice with PBS for 5 min after which they were incubated with blocking buffer [PBS (10% FBS, 0.05% Triton-X)] for 1 h. After blocking, cells were incubated with primary antibody in blocking buffer for 1 h, washed, and incubated with secondary antibody in blocking buffer for 1 h. All labeling steps were performed at room temperature. Nuclei were stained with 14.3 μM DAPI (Thermo Fisher Scientific, MA, USA). Primary antibodies used were: Serca2 ATPase (mouse monoclonal, ab2861, Abcam, UK) (1:500 dilution), Ryanodine receptor (mouse monoclonal, ab2827, Abcam, UK) (1:100), IP3 receptor type 1 (rabbit polycolonal, ab111087, Abcam, UK) (1:100) and Sodium/calcium exchanger (mouse monoclonal, MA3-926, Thermo Fisher Scientific, MA, USA) (1:100). Secondary antibodies were, anti-Mouse IgG (goat polyclonal, A11001, Thermo Fisher Scientific, MA, USA) (1:750) and anti-Rabbit IgG (goat polyclonal, A21245, Thermo Fisher Scientific, MA, USA) (1:750).
Koivumäki J.T., Naumenko N., Tuomainen T., Takalo J., Oksanen M., Puttonen K.A., Lehtonen Š., Kuusisto J., Laakso M., Koistinaho J, & Tavi P. (2018). Structural Immaturity of Human iPSC-Derived Cardiomyocytes: In Silico Investigation of Effects on Function and Disease Modeling. Frontiers in Physiology, 9, 80.
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4

Antibody Characterization for Protein Expression

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Rabbit polyclonal anti-Akt antibody (1:1000 for IB, 9272; Cell Signaling Technology), rabbit polyclonal anti-pAkt-Ser473 antibody (1:1000 for IB, 9271; Cell Signaling Technology), rabbit polyclonal anti-α-tubulin antibody (1:1000 for IB, 2144, Cell Signaling Technology), mouse monoclonal anti-α-actinin antibody (1:400 for IF, A7811; Sigma-Aldrich), mouse monoclonal anti-ryanodine receptor antibody (1:100 for IF, ab2827; Abcam), mouse monoclonal anti-dihydropyridine receptor (DHPR) antibody (1:800 for IF, ab2864; Abcam), mouse monoclonal anti-sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) 2a antibody (1:200 for IF, MA3-919; Thermo-Fisher), rabbit polyclonal anti-STIM1 antibody (1:200, PA1-46217; Thermo-Fisher), mouse monoclonal PLN [2D12] antibody (1:500 for IF, 1:1000 for WB; ab2865, Abcam), mouse monoclonal FLAG antibody (1:500 for IF, 1:1000 for WB, F1804; Millipore-Sigma), mouse monoclonal anti-Reep5 antibody (1:500 for IF, 1:1000 for WB, 14643-1-AP; Proteintech), and rabbit monoclonal anti-KDEL antibody (1:250 for IF, ab176333; Abcam) were used in this study. Goat anti-rabbit Alexa Fluor 488 secondary antibodies (nos. A-11034 and A-11011; Molecular Probes) were used at 1:800 dilution.
Callaghan N.I., Lee S.H., Hadipour-Lakmehsari S., Lee X.A., Ahsan Siraj M., Driouchi A., Yip C.M., Husain M., Simmons C.A, & Gramolini A.O. (2020). Functional culture and in vitro genetic and small-molecule manipulation of adult mouse cardiomyocytes. Communications Biology, 3, 229.
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5

Analyzing Cardiac Protein Expression

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Protein lysates from mouse heart tissues were harvested in radioimmunoprecipitation assay buffer (RIPA, 50 mM Tris-HCl; pH7.4, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, 150 mM NaCl, 2 mM EDTA), supplemented with protease and phosphatase inhibitors (Roche), for 30 mins on ice, spun down at 15,000 × g at 4 °C. Soluble fractions were saved for immunoblotting. Protein lysates were heated for 5 min to either 65 °C or to boiling, run on 4–12% polyacrylamide gels and transferred onto 0.22–0.45 μm nitrocellulose membranes. After blocking for 1 hour with 5% milk in 0.05% TBS-Tween20, primary antibodies were added and incubated at 4 °C overnight: primary mouse monoclonal anti-DHPR (1:500 dilution; ab2864; Abcam), primary mouse monoclonal anti-RyR2 (1:1000 dilution; ab2827; Abcam), primary mouse monoclonal anti-PLN (1:1000 dilution; MA3-922; ThermoFisher), primary mouse monoclonal anti-SERCA2A (1:1000 dilution; MA3-919; ThermoFisher), and primary rabbit polyclonal anti-NCX1 (1:1000 dilution; ab151608; Abcam).
Hadipour-Lakmehsari S., Driouchi A., Lee S.H., Kuzmanov U., Callaghan N.I., Heximer S.P., Simmons C.A., Yip C.M, & Gramolini A.O. (2019). Nanoscale reorganization of sarcoplasmic reticulum in pressure-overload cardiac hypertrophy visualized by dSTORM. Scientific Reports, 9, 7867.
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6

Comprehensive Cardiac Protein Analysis

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Rabbit-Cav1.2 (ACC-003; Alomone Labs), rabbit-RyR2 (MA3-916; Thermo Fisher Scientific), mouse-RyR2 (ab2827; Abcam), mouse-SERCA (NB300-581; Novus Biological), rabbit-ser16-PLN (A010-12; Badrilla), rabbit-thr16-PLN (A010-13; Badrilla), mouse-PLN (A010-14; Badrilla), rabbit-HIF1α (NB100-479; Novus Biological), mouse-HIF2α (clone-190b, MilliporeSigma), rabbit-H3K36me3 (ab9050; Abcam), rabbit-H3K9me3 (ab8898; Abcam), rabbit-H3K4me3 (ab8580; Abcam), rabbit-H3 (ab1791; Abcam), mouse-actin (NB100-74340; Novus Biological), goat anti–rabbit-HRP (sc-2030; Santa Cruz Biotechnology Inc.), goat anti–mouse-HRP (sc-2031; Santa Cruz Biotechnology Inc.), goat anti–rabbit-AlexaFluor 488 (ab15007; Abcam), and goat anti–mouse AlexaFluor 594 (11032; Invitrogen).
Chung Y.J., Luo A., Park K.C., Loonat A.A., Lakhal-Littleton S., Robbins P.A, & Swietach P. (2019). Iron-deficiency anemia reduces cardiac contraction by downregulating RyR2 channels and suppressing SERCA pump activity. JCI Insight, 4(7), e125618.
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