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Anti eea1 antibody

Manufactured by Abcam
Sourced in United States

Anti-EEA1 antibody is a protein that binds to the Early Endosome Antigen 1 (EEA1) protein. EEA1 is a marker for early endosomes and is involved in endocytic membrane fusion processes.

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3 protocols using anti eea1 antibody

1

Endocytosis of Silver Nanoparticles

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To check that the Ag NPs are endocyted by the cells, a FITC conjugation was realized thanks to NH2 groups associated to NPs. (Figure S1). On milliliter of the Ag NPs suspension was added to a solution of FITC in 0.1 M carbonate–bicarbonate buffer and vortexed for 1 minute, and the vial reaction was covered with aluminum foil and incubated for 2 hours at room temperature. To finish, the Ag NPs were washed with cold PBS. The cell lines were incubated with conjugated FITC Ag NPs for 15 minutes. After the incubation, the cells were washed with cold PBS and fixed as described previously, followed by a fluorescent identification of the early endosome marker EEA1. The cells were incubated for 30 minutes with blocking reagent, then incubated overnight with anti-EEA1 antibody (Abcam), and finally for 1 hour with alexa fluor 568 conjugated secondary antibody (Figure S2). The slides were mounted using mounting medium with DAPI (Vector, Cambridgeshire, UK). Finally, the analysis was performed with confocal microscope (Zeiss LSM710; Carl Zeiss Meditec AG, Jena, Germany).
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2

Cellular Uptake of Fluorescent Vaccine Particles

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SAEC and RPMI2650 cells were seeded in 8-well chamber slides at a density of 3×104 cells per well and cultured overnight. Fluorescent vaccine particles were prepared using Cy5 labeled CpG ODN, and then incubated with cells at the ratio of 10–1 between mPSM to cells for 6 h. After incubation, cells were washed with PBS twice, fixed with 4% paraformaldehyde at room temperature for 15 min, and permeabilized with 0.1% tween-20 for 15 min. After blocking with 1% BSA plus 5% FBS, cells were incubated with anti-EEA1 antibody (1:500, Abcam) at 4°C overnight, followed by staining with AF488 -labeled goat anti-rabbit secondary antibody (1:1000 dilution, ThermoFisher) at room temperature for 2 h. Finally, nuclei were stained with 0.5 µg/mL DAPI for 15 min.
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3

Cadmium-Induced Apoptosis Mechanism

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Fetal bovine serum was obtained from Solarbio Company (Beijing, China), and cadmium chloride (CdCl2, purity of ≥ 99.99%, USA) was obtained from Sigma company (St. Louis, MO, USA). Phorbol 12-myristate 13-acetate (TPA, P167764) and nocodazole (N129755) were obtained from Aladdin company (Shanghai, China), and Hochest 33258 (23491-44-3) was obtained from MedChemExpress company (New Jersey, USA). Anti-α-tubulin-FITC antibody and anti-LAMP2 antibody were obtained from Sigma, the monoclonal anti-connexin 43 antibody (CX-1B1, 13-8300) was obtained from Invitrogen company (Waltham, MA, USA), anti-EEA1 antibody, anti-bax (ab32503) antibody, and anti-bcl-2 (ab196495) antibody were obtained from Abcam company (Cambridge, UK), anti-RAB7A antibody was obtained from ABclonal company (Wuhan, China), anti-phospho-connexin 43/GJA1 (Ser373; AF8264) was obtained from Affinity Biosciences (Cincinnati, OH, USA), and mouse anti-active caspase-3 (bsm-33199M) was obtained from BIOSS company (Beijing, China).
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