Ab27043

Manufactured by Abcam

Sourced in United States

Ab27043 is a primary antibody product from Abcam. It is designed for use in various laboratory techniques.

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Lab products found in correlation

7 protocols using ab27043

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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Immunofluorescent Imaging of Cellular Organelles

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Samples were washed with phosphate buffer saline (PBS), fixed with 4% paraformaldehyde (Fluka, Sigma Aldrich) in PBS (w%) for 15 min, permeabilized with 0.5% Triton X-100 (Fluka) in PBS (v%) for 10 min at room temperature and washed with PBS. Samples were then incubated for 1 h with 1% bovine serum albumin (Sigma Aldrich) in PBS (w%). Cells were incubated overnight with an antibody that marks the Golgi apparatus (1:200) produced in mouse and specific for the 58 kDa Golgi protein formiminotransferase cyclodeaminase (ab27043, Abcam, Cambridge, UK), and with TRITC-phalloidin (1:1000) for 10 min, (Sigma Aldrich.) Samples were then stained with Hoechst 34580 (10 μg/mL) for 5 min at room temperature, followed by two rinses of PBS. The fluorescently stained samples were visualized on a Nikon Ti microscope using 20× or 40× air objectives, and imaged using an Andor Neo camera.

Ostrowski M.A., Huang E.Y., Surya V.N., Poplawski C., Barakat J.M., Lin G.L., Fuller G.G, & Dunn A.R. (2015). Multiplexed Fluid Flow Device to Study Cellular Response to Tunable Shear Stress Gradients. Annals of biomedical engineering, 44(7), 2261-2272.

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Immunofluorescence Microscopy of Cellular Proteins

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For immunofluorescence, cells were grown on 12-mm coverslips and fixed with 3% paraformaldehyde (5 min, RT), followed by permeabilization with 0.5% Triton X-100 for 3 min (RT). Further steps were done as described (Hussain et al. 2013 (link)). Imaging was done by confocal laser scanning microscopy (Leica TCS-SP5). Images were processed using TCS-SP5 software. Antibodies against the following proteins were used: CDK5RAP2 (pAb, 06-1398, Millipore), anti-Myc (mAb 9E10), β-actin (mAb A2228, Sigma), α-tubulin (mAb YL1/2), γ-tubulin (mAb, T6557, Sigma), YAP/TAZ (mAb, 8418, Cell Signaling), MST1 (pAb, 3682, Cell Signaling), phospho-MST1/MST2 (pAb, 3681, Cell Signaling), GM130 (mAb, 618822, abcam), 58 K (mAb, ab27043, Abcam), pericentrin (pAb, ab4448, Abcam), γ-tubulin (mAb GTU-88, Sigma), Nesprin-1 (SpecII; (Taranum et al. 2012 (link)), BIRC5 (mAb, ab76424, Abcam), Ki67 as cell proliferation marker (mAb, M 7249, DAKO), anti-PH3 rabbit polyclonal IgG to stain mitotic cells (06-570, Upstate Biotechnology) and Cleaved Caspase-3 (ASP175)-specific polyclonal antibodies as apoptosis marker (No 9661, Cell signaling).

Sukumaran S.K., Stumpf M., Salamon S., Ahmad I., Bhattacharya K., Fischer S., Müller R., Altmüller J., Budde B., Thiele H., Tariq M., Malik N.A., Nürnberg P., Baig S.M., Hussain M.S, & Noegel A.A. (2016). CDK5RAP2 interaction with components of the Hippo signaling pathway may play a role in primary microcephaly. Molecular Genetics and Genomics, 292(2), 365-383.

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Comprehensive Antibody Toolkit for Immunological Research

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The primary antibodies for PD-L1 (13684 and 29122, CST), PD-L1 (17 952-1-AP, Proteintech), PD-L1 (329708, BioLegend), GAPDH (KC-5G5, KANGCHEN), MHC-I (ab70328, Abcam), MHC-I (15 240-1-AP, Proteintech), THADA (ab122260, Abcam), THADA (12 909-1-AP, Proteintech), Sec24A (ab102660, Abcam), Sec24A (15 958-1-AP, Proteintech), Flag-tag (2368, CST), HA-tag (3724, CST), Myc-tag (2276, CST), XTP3-B/ERLEC1 (ab181166, Abcam), OS-9 (ab109510, Abcam), HRD1/SYVN1 (13 473-1-AP, Proteintech), 58K (ab27043, Abcam), GRP94 (ab3674, Abcam), CD8a (D4W2Z; CST), CD8a (CL488-65069; Proteintech), ubiquitin (3936, CST), granzyme B (17215, CST), sodium potassium ATPase (ab76020, Abcam) were all commercially available. Reagents involving recombinant human interferon (IFN)-γ protein (285-IF, R&D Systems), rhPD-1/FC chimera protein (1086-PD-050, R&D Systems), recombinant human IL-2 (589102, BioLegend), cycloheximide (C1988, Sigma-Aldrich), MG132 (M7449, Sigma-Aldrich), chloroquine (CQ) (C6628, Sigma-Aldrich), eeyarestatin I (Eer I) (10012609, Cayman) and 4′,6-diamidino-2-phenylindole (DAPI) (0100-20; SouthernBiotech) were also purchased from the indicated suppliers.

Li C., Chi H., Deng S., Xu K., Wang H., Yao H., Wang Y., Chen D., Guo X., Fang J.Y., He F, & Xu J. (2021). THADA drives Golgi residency and upregulation of PD-L1 in cancer cells and provides promising target for immunotherapy. Journal for Immunotherapy of Cancer, 9(8), e002443.

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Multicolor Fluorescent Labeling Protocol

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Primary antibodies anti-EEA1 (ab70521), anti-58K Golgi (ab27043), antipericentrin (ab28144), and anticatalase (ab110292) were purchased from Abcam, anti-TGN46-8 (SAB4200355) from Sigma and anti-Tom20 (sc-17764) from Santa Cruz Biotechnology. All primary antibodies were monoclonal mouse antibodies. Polyclonal secondary antimouse antibodies conjugated to Atto647N (50185, Sigma Aldrich) as well as isotype-specific secondary antimouse antibodies conjugated to Atto647N (610-156-041, Rockland Immunochemicals) and Alexa Fluor 568 (A-21124, Thermo Fisher Scientific) were used for detection. The nanoboosters—single-domain alpaca antibody fragments covalently coupled to fluorescent dyes—GFP-Booster Atto488 and RFP-Booster Atto594 (Chromotek) were used to boost eYFP and mCherry fluorescence, respectively. Live cell stains SiR-tubulin, SiR-actin, and SiR-lysosome were from Spirochrome and MitoTracker Deep Red FM was from Thermo Fisher Scientific.

Scherer K.M., Manton J.D., Soh T.K., Mascheroni L., Connor V., Crump C.M, & Kaminski C.F. (2021). A fluorescent reporter system enables spatiotemporal analysis of host cell modification during herpes simplex virus-1 replication. The Journal of Biological Chemistry, 296, 100236.

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Immunofluorescence and Cell Culture Analysis

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Immunofluorescence and cell culture analyses were performed as previously described (Schmidts et al., 2015). Briefly, IMCD3, hTERT‐RPE, SH‐SY5Y cells and patient as well as control fibroblasts were cultured in DMEM‐F12 medium with 5% FBS under standard conditions. For ciliation experiments, cells were serum starved for 24 hr. Primary antibodies used: Golgi marker GM130 (mouse, ab169276, Abcam, USA), Golgi marker 58K (ab27043 mouse, Abcam, USA), early endosome marker EEA1 (1G11 ab70521, mouse monoclonal, Abcam), PPP1R21 antibody (rabbit, HHPA036792 (ab1) and HPA 036791 (ab2), Atlas antibodies, Sweden), acetylated tubulin (mouse monoclonal 6‐11‐B1, Sigma, USA). In brief, cells were grown on coverslips, washed with PBS and fixed 5 min in 5% PFA. After three washes, cells were permeabilized using 0.1% Triton for 3 min, washed three times and incubated in 5% BSA for 1 hr at RT to bock unspecific reactions. Incubation in primary antibodies was then performed for 1 hr (1:200 in 5% BSA), after five washes, cells were incubated in corresponding fluorescence tagged secondary 1:1,000 antibody dilutions for 30 min, washed five times in PBS and then mounted in Vectashield with DAPI. Images were taken with a Zeiss Apotome Axiovert 200 and processed with AxioVision 4.8 and Fiji software.

Rehman A.U., Najafi M., Kambouris M., Al‐Gazali L., Makrythanasis P., Rad A., Maroofian R., Rajab A., Stark Z., Hunter J.V., Bakey Z., Tokita M.J., He W., Vetrini F., Petersen A., Santoni F.A., Hamamy H., Wu K., Al‐Jasmi F., Helmstädter M., Arnold S.J., Xia F., Richmond C., Liu P., Karimiani E.G., Karami Madani G., Lunke S., El‐Shanti H., Eng C.M., Antonarakis S.E., Hertecant J., Walkiewicz M., Yang Y, & Schmidts M. (2018). Biallelic loss of function variants in PPP1R21 cause a neurodevelopmental syndrome with impaired endocytic function. Human Mutation, 40(3), 267-280.

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Immunofluorescence Labeling of Cellular Junctions

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The following primary antibodies were used: goat polyclonal anti‐VE‐cadherin (1:200, sc‐6458, Santa Cruz Biotechnology), anti‐β‐catenin (1:100, #610 154, BD Transduction Laboratories), rabbit polyclonal anti‐pY658‐VEC^[¹⁴^] (1µg mL⁻¹), and mouse monoclonal anti‐58K Golgi protein (1:100, ab27043, Abcam). The secondary antibodies were donkey anti‐goat‐Alexa 488 (Invitrogen, A‐11055), donkey anti‐Mouse‐Alexa 555 (Invitrogen, A‐31570), chicken anti‐rabbit‐Alexa 647 (Invitrogen, A‐21443), and donkey anti‐mouse‐Alexa 488 (Invitrogen, #A21202).

Stefopoulos G., Lendenmann T., Schutzius T.M., Giampietro C., Roy T., Chala N., Giavazzi F., Cerbino R., Poulikakos D, & Ferrari A. (2022). Bistability of Dielectrically Anisotropic Nematic Crystals and the Adaptation of Endothelial Collectives to Stress Fields. Advanced Science, 9(16), 2102148.

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