Ab11826

Manufactured by Abcam

Sourced in United States

Ab11826 is a polyclonal primary antibody produced in rabbit. It is designed to detect the target protein in various applications. The core function of this product is to specifically bind and detect the target protein.

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Lab products found in correlation

32 protocols using ab11826

ChIP-seq protocol for histone modifications

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ChIP assay was conducted as described previously in^{50 (link)}. In brief, HSV-1 infected cells were fixed with 1% formaldehyde and sonicated to shear DNA. After centrifugation, the supernatants were incubated with anti-SRSF2 antibodies (Abcam, ab11826), anti-H3K4Me3 antibodies (Abcam, ab8580), anti-H3K27Me3 antibodies (Abcam, ab6002), anti-H3K27Ac antibodies (Abcam, ab4729), or anti-histone H3 antibodies (Abcam, ab1791). Chromatin DNA was purified by Dynabeads protein G (Invitrogen, 10004D) and subjected to real-time PCR. The region-specific primers are listed in Supplementary Table 1.

Li K, & Wang Z. (2021). Speckles and paraspeckles coordinate to regulate HSV-1 genes transcription. Communications Biology, 4, 1207.

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Immunoprecipitation Assay with Dynabeads

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The immunoprecipitation assay was performed using the Immunoprecipitation Protein G Dynabeads^® kit (Invitrogen, 10007D) according to the manufacturer’s protocol. The antibodies used for immunoprecipitation included anti-SRSF2 antibody (Abcam, ab11826), anti-P300 antibody (Abcam, ab59240), anti-CBP antibody (Abcam, ab50702), anti-STAT3Y705p antibody (Abcam, ab76315), and anti-H3K27AC antibody (Abcam, ab4729).

Wang Z., Li K., Chen W., Wang X., Huang Y., Wang W., Wu W., Cai Z, & Huang W. (2019). Modulation of SRSF2 expression reverses the exhaustion of TILs via the epigenetic regulation of immune checkpoint molecules. Cellular and Molecular Life Sciences, 77(17), 3441-3452.

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Culturing and Immunostaining HeLa Cells

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HeLa cells (ATCC^® CCL-2) were cultured in DMEM with high glucose, 10% fetal bovine serum, 100 U/mL penicillin and 100 mg/mL streptomycin (all from Millipore-Sigma, Burlington, MA, USA), in a humidified incubator with 5% CO₂ at 37 °C. For peptide treatment, 5 × 10⁵ cells were seeded in 6-well plates with 2 mL of full-DMEM medium 24 h before the treatment. Next day, the medium was exchanged with pre-warmed DMEM medium supplemented with respective peptides and incubated for defined times. As a negative control, HeLa cells were incubated with pre-warmed DMEM medium supplemented with 1× PBS without any of the CPPs.
Antibodies against fibrillarin (ab4566), coilin (ab87913), Sc35 (ab11826), tubulin (ab195883), 58K (ab27043) and M6PR (ab124767) were all purchased from Abcam (Cambridge, MA, USA). Alexa Fluor 555 Goat Anti-Rabbit (ab150078) and Alexa Fluor 488 Goat Anti-Mouse (ab150117), used as secondary antibodies, were also purchased from Abcam. Hoechst 43222 (H1399) were purchased from Invitrogen (Waltham, MA, USA).

Venit T., Dowaidar M., Gestin M., Mahmood S.R., Langel Ü, & Percipalle P. (2020). Transcriptional Profiling Reveals Ribosome Biogenesis, Microtubule Dynamics and Expression of Specific lncRNAs to be Part of a Common Response to Cell-Penetrating Peptides. Biomolecules, 10(11), 1567.

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Immunofluorescence and Cell Cycle Analysis

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Mouse anti-SC35 antibody (1∶250, Abcam, ab11826) or mouse anti-coilin antibody (1∶50, Abcam, ab87913) were used for the protein detections^{40 (link)}. In this case, a particular antibody was added to staining solution. After the washing and rinsing step, proteins were detected using Alexa Fluor 488 anti-mouse antibody (1∶100) diluted in 25 mM Tris-HCl, pH 7.4, 150 mM NaCl. Protein detection was evaluated prior to the elution step.
DNA replication was detected using EdU. The cells were incubated with 10 µM EdU for 30 minutes prior to the ethanol fixation. Then, EdU was detected using a click reaction^{36 (link)} followed by the developed approach. The EdU signal was analysed prior to the elution step.
If the cell cycle analysis was performed, the image cytometry was performed prior to the elution step. The cell cycle analysis by image cytometry was performed according to^{37 (link)}.

Ligasová A, & Koberna K. (2019). Quantification of fixed adherent cells using a strong enhancer of the fluorescence of DNA dyes. Scientific Reports, 9, 8701.

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Ultrastructural and Protein Localization

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The experiments are performed as our previous described procedures. Thirty-nine EM images were obtained from thin sections using a JEM1400 electron microscope (JEOL, Akishima, Tokyo, Japan). The relative cytosolic areas in cross-sections of 10 cells were analyzed by Image-Pro Plus software. And fluorescence signals obtained using anti-FLAG (mouse) (A8592, Sigma), anti-FLAG (rabbit) (20543-1-AP, Proteintech), anti-HA (mouse) (H3663, Sigma), anti-HA (rabbit) (C29F4, Cellular signaling), anti-SC35 antibody (Ab11826, Abcam), and anti-SF2 antibody (sc-33652, Santa Cruz) were acquired and analyzed by Carl Zeiss LSM 880 microscope (Carl Zeiss, Germany). At least 10 cells from each group were analyzed.

Chen Z.H., Chen T.Q., Zeng Z.C., Wang D., Han C., Sun Y.M., Huang W., Sun L.Y., Fang K., Chen Y.Q., Luo X.Q, & Wang W.T. (2020). Nuclear export of chimeric mRNAs depends on an lncRNA-triggered autoregulatory loop in blood malignancies. Cell Death & Disease, 11(7), 566.

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Immunofluorescence Staining of Nuclear Bodies

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The following antibodies were used: rabbit anti-Coilin (1:2000; ab210785, Abcam), mouse IgG1 anti-Fibrillarin (1:60; ab4566, Abcam), mouse IgG1 anti-nuclear pore complex proteins MAb414 (1:1000; MMS-120P-100, Eurogentec), rabbit anti-NPAT (1:700; A302–772A, Bethyl Laboratories), mouse IgG1 anti-PSPC1(1:100; clone IL4, SAB4200503, Sigma-Aldrich), rabbit anti-pSF3b155 (phosphorylated at Thr313; 1:400; clone D8D8V, 25009, Cell Signaling), rabbit anti-SF3b155 (1:600; clone D7L5T, 14434, Cell Signaling), mouse IgG1 anti-SMN1 (1:100; Survival of Motor Neurons, clone 2B1, 05–1532, EMD Millipore), mouse IgG1 anti-SRSF2/SC35 (1:400; ab11826, Abcam), mouse IgG1 anti-TDP-43/TARDBP (1:1500; clone 3H8, MABN45, EMD Millipore), and species-specific Alexa Fluor secondary antibodies (1:400; Thermo Fisher). A recent study^{45 (link)} proposed that the main target of the monoclonal SRSF2/SC35 antibodies is SRRM2 instead of SRSF2/SC35. However, SRRM2 is a spliceosome-associated protein that sharply localizes to nuclear speckles^{45 (link)}. We also confirmed that the monoclonal ab11826 antibody we used recognized SRS2-GFP⁺ speckles in fixed oocytes expressing SRSF2-GFP. The conclusions of our study are thus unaffected by the proposed SRSF2/SC35 antibody discrepancy^{45 (link)}.

Al Jord A., Letort G., Chanet S., Tsai F.C., Antoniewski C., Eichmuller A., Da Silva C., Huynh J.R., Gov N.S., Voituriez R., Terret M.É, & Verlhac M.H. (2022). Cytoplasmic forces functionally reorganize nuclear condensates in oocytes. Nature Communications, 13, 5070.

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Immunofluorescence Assay for HeLa Cells

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HeLa cells were seeded on coverslips and transfected with 0.25 μg of plasmid DNA. Cells were fixed in 4% paraformaldehyde for 10 min and permeabilized with 0.1% Triton X-100 in PBST (PBS + 0.01% Tween 20) for 10 min at room temperature. The cells on coverslips were blocked with blocking solution (2% BSA/PBST) for 30 min and incubated with the primary antibodies anti-Flag (mouse, Sigma, M2, F1804), anti-PABPN1 (rabbit, Abcam, ab75855), anti- SC35 (mouse, Abcam, ab11826) diluted in the blocking solution overnight at 4 °C. Proteins were visualized with the FITC- and TRITC-conjugated secondary antibodies (Sigma, F6005, and T5393) for 1 h at room temperature. Nuclei were stained with DAPI-supplemented Fluoromount-G mounting media (Thermo Fisher Scientific). Cells were visualized with a fluorescence microscope (Nikon eclipse 90i), and the images were analyzed with the NIS-elements (Nikon) software.

Kases K., Schubert E., Hajikhezri Z., Larsson M., Devi P., Darweesh M., Andersson L., Akusjärvi G., Punga T, & Younis S. (2023). The RNA-binding protein ZC3H11A interacts with the nuclear poly(A)-binding protein PABPN1 and alters polyadenylation of viral transcripts. The Journal of Biological Chemistry, 299(8), 104959.

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Immunofluorescence staining of RNA methylation proteins

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The cells were seeded onto the sterile slides and placed in the 6 well plates with the cell density of 1 × 10⁵ cells per well. After incubation for 24h, the cells were gently rinsed by phosphate-buffered saline (PBS) 3 times followed by fixed with 4% paraformaldehyde for 15min. Then, the cells were washed with PBS three times followed by permeabilized by 0.1% Triton X-100 in PBS for 15 min. To block the nonspecific binding, the cells were incubated with 2% bovine serum albumin (BSA) in PBS for 1h. The cells were subsequently incubated with primary antibodies against METTL16 (1:100, HPA020352, Sigma-Aldrich), METTL3 (1:100, ab195352, Abcam), METTL 14 (1:100, HPA038002, Sigma-Aldrich) and SC35 (1:500, ab11826, Abcam) overnight at 4°C, respectively. After washing with PBS three times, the corresponding fluorescence-labeled secondary antibodies (1:200, goat anti-rabbit IgG(H+L) Alexa Fluor 555: 4413S, Cell Signaling Technology; 1:200, goat anti-mouse IgG(H+L) Alexa Fluor 488: 4408S, Cell Signaling Technology) were applied to stain the cells and kept incubating for 1h. The nuclei were counterstained by mounting the cells in DAPI (F6057, Sigma-Aldrich). The immunofluorescent signals were photographed by a confocal laser scanning microscope (CLSM) (Carl Zeiss LSM 880).

Su R., Dong L., Li Y., Gao M., He P.C., Liu W., Wei J., Zhao Z., Gao L., Han L., Deng X., Li C., Prince E., Tan B., Qing Y., Qin X., Shen C., Xue M., Zhou K., Chen Z., Xue J., Li W., Qin H., Wu X., Sun M., Nam Y., Chen C.W., Huang W., Horne D., Rosen S.T., He C, & Chen J. (2022). METTL16 exerts an m6A-independent function to facilitate translation and tumorigenesis. Nature cell biology, 24(2), 205-216.

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Immunofluorescence Analysis of SF3B1 and SC35

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One million cells of indicated genotypes were seeded onto Corning BioCoat Fibronectin 22 mm cover-slips (Fisher Scientific 08-774-386) in six-well plates. After 2 days, cells were fixed with 4% paraformaldehyde/phosphate-buffered saline (PBS) for 20 min at room temperature (RT). After 3 × PBS wash, cells were permeabilized with 0.1% Triton X-100/PBS for 20 min at RT. After 3 × PBS wash, cells were blocked with 5% FBS/PBS for 1 h at RT and incubated with α-SF3B1 mouse monoclonal antibody (MBL, D221-3) or α-SC35 mouse monoclonal antibody (Abcam, ab11826) at 1:50 dilution in 5% FBS/PBS in cold room overnight. On the second day, coverslips were washed with PBS three times and incubated with Alexa Fluor 488 anti-mouse secondary antibody (Thermo Fisher Cat #: A-11029) at 1:500 dilution in 5% FBS/PBS at RT in dark for 1 h. Coverslips were then washed with PBS three times and mounted using ProLong Gold Antifade Mountant with DAPI (Thermo Fisher, P36935). Slides were imaged with a × 40 objective on an Olympus IX-81 inverted fluorescence microscope and imaged, captured and processed with Metamorph for Olympus.

Teng T., Tsai J.H., Puyang X., Seiler M., Peng S., Prajapati S., Aird D., Buonamici S., Caleb B., Chan B., Corson L., Feala J., Fekkes P., Gerard B., Karr C., Korpal M., Liu X., T. Lowe J., Mizui Y., Palacino J., Park E., Smith P.G., Subramanian V., Wu Z.J., Zou J., Yu L., Chicas A., Warmuth M., Larsen N, & Zhu P. (2017). Splicing modulators act at the branch point adenosine binding pocket defined by the PHF5A–SF3b complex. Nature Communications, 8, 15522.

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Immunofluorescence Analysis of TDP-43, SRSF1, SRSF2, and NCL

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Slides with NSC34 cells were fixed and permeabilised in 4% PFA/0.2% Triton X-100 at RT for 20 min. Slides were incubated with rabbit polyclonal anti-TDP-43 (1 in 200, Proteintech, 10782-2-AP), rabbit polyclonal anti-SRSF1 (1 in 200, Abcam ab38017), mouse monoclonal anti-SRSF2 (1 in 200, Abcam ab11826), or rabbit polyclonal anti-NCL (1:200; Proteintech; 10556-1-AP) in 2% BSA in PBS at RT for 1 h. Slides were washed three times in PBS, then incubated with goat anti-rabbit IgG H&L (1 in 1000; Alexa Fluor® 488; Abcam) or goat anti-mouse IgG H&L (1 in 1000; Alexa Fluor® 488; Abcam) in 2% BSA in PBS at RT for 1 h. Formalin fixed paraffin-embedded tissue sections were deparaffinized and mouse anti-NCL (1 in 150 in 5% BSA; Abcam; ab136649) stained slides were first antigen retrieved by heat in trisodium citrate pH 6.5 for 20 min, then completed as for ICC. Slides of NSC34 and tissue sections were mounted with mounting medium containing DAPI (Vector Labs Inc.).

Stopford M.J., Higginbottom A., Hautbergue G.M., Cooper-Knock J., Mulcahy P.J., De Vos K.J., Renton A.E., Pliner H., Calvo A., Chio A., Traynor B.J., Azzouz M., Heath P.R., Kirby J, & Shaw P.J. (2017). C9ORF72 hexanucleotide repeat exerts toxicity in a stable, inducible motor neuronal cell model, which is rescued by partial depletion of Pten. Human Molecular Genetics, 26(6), 1133-1145.

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