For imaging flow cytometry experiments: anti-CD8 APC-Cy7 (BD Biosciences, San Jose, CA or Tonbo Biosciences, San Diego, CA) and anti-CD56 PE-CF594 (BD Biosciences) were used for surface staining; all intracellular stains included anti-perforin D48 PE or BV421 (Biolegend, San Diego, CA) and anti-perforin δG9 FITC (Biolegend) and one of the following unconjugated antibodies: anti-CD71 (Cell Signaling Technology, Danvers, MA), anti-granzyme B (BD Biosciences), anti-rab3D (Abcam, Cambridge, MA), anti-rab4 (Pierce Antibodies, Waltham, MA), anti-rab5 (Cell Signaling Technology), anti-rab7 (Santa Cruz Biotechnology, Dallas, TX), anti-rab8 (Cell Signaling Technology), anti-rab11a (Cell Signaling Technology), anti-rab27a (Abcam), anti-rab35 (Abcam), anti-rab37 (Abcam), anti-syntaxin6 (Cell Signaling Technology), anti-syntaxin7 (R&D systems, Minneapolis, MN), anti-vti1b (Abcam), anti-VAMP3 (Abcam), anti-VAMP4 (Abcam), or anti-SNAP23 (Abcam). The unconjugated antibodies were detected using goat anti-rabbit AF647, chicken anti-goat AF647, or donkey anti-sheep AF647 secondary antibodies (Invitrogen, Carlsbad, CA). For confocal microscopy, perforin D48 FITC (Abcam), perforin δG9 AF647 (Biolegend), and goat anti-rabbit or donkey anti-sheep AF568 secondary antibodies (Invitrogen) were used, along with the selected rab or SNARE antibody.
Anti rab5
Manufactured by Cell Signaling Technology
Sourced in United States
Anti-Rab5 is a primary antibody that specifically recognizes the Rab5 protein. Rab5 is a small GTPase that plays a key role in the regulation of early endocytic vesicle formation and trafficking. The Anti-Rab5 antibody can be used in various experimental techniques, such as Western blotting, immunoprecipitation, and immunocytochemistry, to detect and study the Rab5 protein.
Automatically generated - may contain errors
Lab products found in correlation
Anti rab7, by Cell Signaling Technology (16 mentions)
Anti rab9, by Cell Signaling Technology (5 mentions)
Hoechst 33342, by Thermo Fisher Scientific (5 mentions)
Anti eea1, by Cell Signaling Technology (4 mentions)
Anti lamp1, by Cell Signaling Technology (3 mentions)
Anti rab11, by Cell Signaling Technology (3 mentions)
Anti rab11a, by Cell Signaling Technology (2 mentions)
Anti lamp1, by Abcam (2 mentions)
Anti caveolin 1, by Merck Group (2 mentions)
Anti lamp2, by Abcam (2 mentions)
Anti gm130, by Cell Signaling Technology (2 mentions)
Celltrace violet, by Thermo Fisher Scientific (2 mentions)
Alexa fluor 488 donkey anti rabbit igg, by Thermo Fisher Scientific (2 mentions)
Goat anti human igg alexafluor 546, by Thermo Fisher Scientific (2 mentions)
Anti rab11, by Thermo Fisher Scientific (2 mentions)
Ticilimumab, by Atlas Antibodies (2 mentions)
Anti lrba, by Atlas Antibodies (2 mentions)
Anti kif5b, by Abcam (2 mentions)
Anti erk, by Cell Signaling Technology (2 mentions)
Anti lc3a b, by Cell Signaling Technology (2 mentions)
27 protocols using anti rab5
1
Imaging Flow Cytometry for Immune Cell Analysis
2
Western Blot Analysis of Rab GTPases
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Total cell extracts were used for western blotting as described [35 (link)]. Blots were detected using the antibodies for anti-Rab5 (1:1000; Cell Signaling), anti-Rab7 (1:1000; Cell Signaling), anti-Rab11 (1:1000; BD Transduction, San Jose, CA, USA), and anti-GAPDH (1:10000; Cedarlane Lab, Hornby, ON, Canada). Staining signal was detected with peroxidase-conjugated secondary antibodies and enhanced chemiluminescence detection system.
3
Immunofluorescence Analysis of Organelle Markers
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Cells were fixed with 4% formaldehyde (Thermo Fisher Scientific) in PBS for 15 min at room-temperature. Cells were washed three times with PBS followed by blocking and permeabilization with 1% BSA (Sigma-Aldrich, Saint Louis, Missouri, USA)/0.3% Triton X-100 in PBS for 1 hr. Antibodies were diluted in blocking/permeabilization buffer and incubated for 2 hr at room temperature. Primary antibodies used were: anti-GFP (clone 5G4, Ogris lab, MFPL), anti-GRASP65 (Thermo Fisher, PA3-910), anti-Rab5 (Cell Signaling Technology, #C8B1), anti-Rab7 (Cell Signaling Technology, #D95F2) and anti-LAMP1 (Abcam, Cambridge, UK, #ab24170). Cells were washed three times with PBS-Tween20 (0.05%) for 5 min each, followed by secondary antibody incubation in blocking/permeabilization buffer for 1 hr at room-temperature. Secondary antibodies used were: goat anti-mouse IgG (H + L) Alexa Fluor 488 (Thermo Fisher A11001), goat anti-rabbit IgG (H + L) Alexa Fluor 555 (Thermo Fisher, A21428), Cells were counterstained with DAPI (Thermo Fisher) for 10 min in PBS. Cells were mounted with ProLong Gold Antifade Mountant (Thermo Fisher #P36930). Images were acquired using a Plan-Apochromat 40x/1.4 Oil DIC objective M27 on a Zeiss LSM880 confocal microscope. Pictures were processed with ImageJ.
4
Monoclonal Antibody Characterization of RTX Toxin
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Anti-adenylate cyclase toxin RTX domain mouse monoclonal antibody (MAb 9D4) was from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-Rab-5 and anti-LAMP-1 from Cell Signalling technology (USA); anti-caveolin-1, sucrose, ficoll, methyl-β-cyclodextrin, nystatin, nifedipine, chlorpromazine, and genistein from Sigma-Aldrich (St Louis, MI, USA); KT5720, calpeptin, and Okadaic acid from Calbiochem (Merck, Germany); Fluospheres® sulphate microspheres (latex beads), Hoechst, anti-mouse Texas Red®, anti-rabbit FITC, Vibrant DiI and Alexa Fluor® phalloidin from Invitrogen, Molecular Probes (Carlsbad, CA, USA). BactoTM proteose peptone, DifcoTM Bordet Gengou Agar base from BD Biosciences (Spain). Gentamicin was from Gibco Thermo Fisher Scientifics, USA.
5
Immunofluorescence Imaging of Organelle Markers
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HeLaClover-PRP4K cells were plated onto glass coverslips in a 6-well plate and treated overnight with 50 μm chloroquine (Sigma) or stimulated with EGF as above. Cells were fixed with 4% paraformaldehyde in PBS for 10 min at room temperature, permeabilized with 0.1% Triton X-100 in PBS for 10 min and immunolabeling was carried out as previously described.10 (link) Fluorescent images were captured with a Zeiss Cell Observer spinning-disk microscope (Intelligent Imaging Innovations, 3i, Boulder, CO, USA) using a 1.4 NA 63 × immersion oil objective lens. Images were processed using only linear adjustments in Adobe Photoshop CS5 and Slidebook (3i) software, which was also used for the analysis of mean fluorescence intensity of immunostaining.
Antibodies used for immunofluorescence were used at the manufacturers recommended dilution unless otherwise noted, and include anti-GFP (Abcam, Toronto, ON, Canada, ab13970) (1:2000 dilution), anti-EEA1 (Cell Signaling, #3288) (1:100 dilution), anti-Rab5 (Cell Signaling, #3547) (1:200 dilution), anti-Rab7 (Cell Signaling, #9367) (1:100 dilution), anti-p62 (Cell Signaling, #7695) (1:400 dilution), anti-LAMP2 (Abcam, ab25631) (1:200 dilution), and anti-p-EGFR (Tyr1068) (Cell Signaling, #3777) (1:800 dilution).
Antibodies used for immunofluorescence were used at the manufacturers recommended dilution unless otherwise noted, and include anti-GFP (Abcam, Toronto, ON, Canada, ab13970) (1:2000 dilution), anti-EEA1 (Cell Signaling, #3288) (1:100 dilution), anti-Rab5 (Cell Signaling, #3547) (1:200 dilution), anti-Rab7 (Cell Signaling, #9367) (1:100 dilution), anti-p62 (Cell Signaling, #7695) (1:400 dilution), anti-LAMP2 (Abcam, ab25631) (1:200 dilution), and anti-p-EGFR (Tyr1068) (Cell Signaling, #3777) (1:800 dilution).
6
Comprehensive Alzheimer's Disease Biomarker Assay
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Anti-ALDH2 (Proteintech, 15310-1-AP, 1:4000), anti-4-HNE (Abcam, ab46545, 1:2000), anti-APP C-Terminal Fragment (C1/6.1, Biolegend, 802801, 1:500), anti-β-amyloid (1–16) (6E10, Biolegend, 803001, 1:1000), anti-β-amyloid (1–40) (Cell Signaling Technology, 12990, 1:1000), anti-β-amyloid (1–42) (Cell Signaling Technology, 14974, 1:1000), anti-GM130 (Cell Signaling Technology, 12480S, 1:1000), anti-TGN46 (Invitrogen, MA3-063, 1:1000), anti-presenilin 1 (Sigma, MAB5232, 1:500), anti-PEN2 (Abcam, ab18189, 1:500), anti-APH1 (Invitrogen, PA1-2010, 1:1000), anti-Rab5 (Cell Signaling Technology, 3547, 1:1000), anti-RCAS1 (Cell Signaling Technology, 12290S, 1:1000), anti-VDAC (Cell Signaling Technology, 4661S, 1:1000), anti-LAMP2 (Proteintech, 27823-1-AP, 1:1000), anti-CANX (Cell Signaling Technology, 2679, 1:1000), anti-VPS35 (Abcam, ab10099, 1:1000), anti-LC3A (Cell Signaling Technology, 4599, 1:1000), anti-Iba1 (Wako, 019-19741, 1:500), anti-tau (Cell Signaling Technology, 46687, 1:1000), anti-phospho-S396-tau (Abcam, ab32057, 1:1000), anti-α-tubulin (GeneTex, GTX628802, 1:10,000), anti-β-actin (GeneTex, GTX124213, 1:10,000), anti-GAPDH (MBL, M171-3, 1:10,000), and anti-SorLA/SORL1 antibody (Abcam, ab190684, 1:1000).
7
Immunofluorescence staining of fixed cells
8
Immunoblot analysis of Caveolin proteins
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Immunoblots were analyzed as described elsewhere [21 (link)]. Detergent-soluble protein fractions were prepared from Caco-2 monolayers incubated with TW or controls and detected by the primary monoclonal anti-Caveolin-1 (Sigma-Aldrich), anti-Caveolin 2 (BD Pharmingen, Heidelberg), anti-rab5, anti-rab7, and anti-cathepsin (Cell Signaling, Cambridge), and anti-TW antibodies.
9
Colocalization of APP and Rab GTPases
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Cortical neurons derived from PDAPP Becn1+/+ and PDAPP Becn1FA/FA embryos were grown on poly-L-lysine coated culture slides. Cells (9 DIV) were then fixed in 4% paraformaldehyde and permeabilized with 0.3% Triton X-100. Slides were blocked for 1 h in PBS containing 1% BSA and 2% normal goat serum and then incubated overnight at 4°C with primary antibodies: anti-APP (Biolegend; 803001) and anti-Rab5 (Cell Signaling Technology; 3547) or anti-Rab7 (Cell Signaling Technology; 9367). After washing, slides were incubated with species-specific Alexa-dye conjugated secondary antibodies for 1 h at room temperature. Slides were sealed with coverslip using mounting medium containing DAPI (Vectashield) and then analyzed by confocal microscopy. Confocal images were collected on Nikon A1 microscope using a 60x oil immersion objective lens and NIS Elements software. The Mander’s colocalization coefficient and the fluorescence intensity profile were generated using the NIS Element software.
10
Immunofluorescence Staining of SARS-CoV-2 Proteins
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Primary and secondary antibodies were diluted at 1:500 for immunofluorescence as follows: anti-Rab5 (#3547, Cell Signaling, Danvers, MA, USA), anti-Spike S1 antibody (#100715-2, BPS, San Diego, CA, USA), anti-angiotensin converting enzyme 2 (ACE2, SC390851, Santa Cruz Biotechnology, Dallas, TX, USA), anti-TOMM20 (11802-1-AP, Proteintech, Rosemont, IL, USA), anti-rabbit IgG AF488 (#A32731, Invitrogen, Waltham, MA, USA), anti-mouse IgG AF568 (#A11004, Invitrogen), and anti-human IgG AF488 (#A11013, Invitrogen). The receptor binding domain (RBD, S1) of spike protein (BT10569, R&D Systems, Minneapolis, MN, USA) and active trimer of spike protein (10586-CV, R&D Systems) were purchased from R&D Systems.
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