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Alexa fluor conjugates

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Alexa Fluor conjugates are a series of highly fluorescent dyes developed by Thermo Fisher Scientific. These dyes are designed to be covalently attached to proteins, antibodies, and other biomolecules, enabling them to be used as labels for various biological applications, such as flow cytometry, fluorescence microscopy, and immunoassays. The Alexa Fluor dyes offer a range of excitation and emission wavelengths, allowing for flexibility in experimental design and compatibility with different imaging and detection systems.

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Lab products found in correlation

4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (7 mentions) 4 6 diamidino 2 phenylindole (dapi), by Vector Laboratories (4 mentions) Mouse anti biotin, by Jackson ImmunoResearch (4 mentions) 4 6 diamidino 2 phenylindole (dapi), by Merck Group (3 mentions) Prolong gold, by Thermo Fisher Scientific (3 mentions) Ab93808, by Abcam (3 mentions) Triton x 100, by Merck Group (3 mentions) Sheep anti gfp, by Bio-Rad (3 mentions) Sheep anti digoxigenin dig, by Roche (3 mentions) Filter paper, by Merck Group (2 mentions) Mab5046, by Merck Group (2 mentions) Ab144p, by Merck Group (2 mentions) Eclipse c1, by Nikon (2 mentions) Smi 32r 100, by Fortrea (2 mentions) Alexa fluor conjugated phalloidin, by Thermo Fisher Scientific (2 mentions) Mab386, by Merck Group (2 mentions) Axio imager, by Zeiss (2 mentions) Prolong gold antifade reagent, by Thermo Fisher Scientific (2 mentions) Paraformaldehyde (pfa), by Merck Group (2 mentions) Nocodazole, by Merck Group (2 mentions)

Close spelling variants of this product

Alexa Fluor phalloidin conjugates (2 citations) Alexa Fluor 488 or 555 conjugates (2 citations) Alexa Fluor antibody conjugates (2 citations) Annexin V-Alexa Fluor-488 and -647 conjugates (2 citations) Alexa Fluor conjugates of (3 citations) Alexa Fluor 488 and 555 conjugates (2 citations) Goat anti-Rat IgG (H+L) Alexa Fluor®488 conjugates (2 citations) Alexa Fluor® 488 conjugate of WGA solution (3 citations) Streptavidin Alexa Fluor® 647 Conjugate (5 citations) Streptavidin–Alexa Fluor 633 conjugate (2 citations)

56 protocols using alexa fluor conjugates

1

Histological Analysis of Brain Tissue

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Animals were euthanized by cervical dislocation; brains were removed, and the right hemisphere was immediately fixed in 4% paraformaldehyde (PFA) at 4 °C overnight and embedded in paraffin for histological analyses. Five-micrometer paraffin sagittal sections were obtained by microtome and sections were deparaffinized and hydrated with deionized water. The detection of PGs by periodic acid Schiff (PAS) staining and immunohistochemical analysis were performed as detailed in [29 (link)]. For immunohistochemistry, primary and secondary antibodies used were guinea pig anti-GFAP (1:500, Synaptic Systems #173_004), rabbit anti-Iba1 (1:200, WAKO #019–19741), Alexa Fluor-conjugates [1:500, Life Technologies: goat anti-guinea pig IgG Alexa Fluor® 594 (#A11076)], and goat anti-rabbit IgG [Alexa Fluor® 488 (#A11008)]. Background controls of secondary antibodies were performed in parallel. Nuclear staining was performed with DAPI (Sigma-Aldrich). Coverslips were mounted with Fluoromount-G™ (Thermo Fisher Scientific).
Mollá B., Heredia M., Campos Á, & Sanz P. (2022). Pharmacological Modulation of Glutamatergic and Neuroinflammatory Pathways in a Lafora Disease Mouse Model. Molecular Neurobiology, 59(10), 6018-6032.
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2

Quantitative Immunohistochemical Analysis of Tumor Markers

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Cells were lysed in RIPA buffer (Sigma) with protease inhibitor cocktail
(Roche and phosphatase inhibitor cocktails (Santa Cruz Biotechnology) for
Western blotting. Paraffin-embedded formalin-fixed sections were subjected to
heat-induced antigen retrieval. The biotin-streptavidin system (Vector
Laboratories) and DAB were used to visualize human Ki-67, while Alexa Fluor
conjugates (Life Technologies) and TSA plus fluorescein system (Perkin Elmer)
were used to detect human PDGFRβ, EGFR, c-MET and mouse ki-67, PD-L2, CD8
and CD11c. For immuno-cytochemistry, cells were fixed with 4% PFA and stained
with Alexa Fluor 555 (for Ki-67) and Alexa Fluor 488 (for CD271) conjugates
(Life Technologies). Nuclei were counterstained by hematoxylin or DAPI.
Fluorescent images were digitized using the Applied Imaging Ariol SL-50scanner
at 20x magnification. Quantification of CD8 and Ki-67 signals was performed
using the Definiens Tissue Studio 4.3 (Definiens, Inc.) according to the
manufacturer’s instruction. Please refer to Supplementary Methods for primary
antibody information. Cell surface PD-L1 and PD-L2 were detected by flow
cytometry using APC-conjugated anti-PD-L1 and PE-conjugated anti-PD-L2
antibodies (BioLegend).
Song C., Piva M., Sun L., Hong A., Moriceau G., Kong X., Zhang H., Lomeli S., Qian J., Yu C.C., Damoiseaux R., Kelley M.C., Dahlman K.B., Scumpia P.O., Sosman J.A., Johnson D.B., Ribas A., Hugo W, & Lo R.S. (2017). Recurrent tumor cell-intrinsic and -extrinsic alterations during MAPKi-induced melanoma regression and early adaptation. Cancer discovery, 7(11), 1248-1265.
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3

Immunolabeling of Retinal Cell Types

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Following Ca2+ imaging, retinas were mounted onto filter paper (0.8 μm pore size, Millipore) and fixed in 4% paraformaldehyde (in PBS) for 15 min at 4°C. Immunolabelling was performed using antibodies against ChAT (choline-acetyltransferase; goat anti-ChAT, 1:100, AB144P, Millipore), GAD67 (glutamate decarboxylase; mouse anti-GAD67, 1:100, MAB5046, Millipore), SMI-32 (mouse anti-SMI32, 1:100, SMI-32R-100, Covance), and melanopsin (rabbit anti-melanopsin, 1:4000, AB-N38, Advanced Targeting Systems) for 4 days. Secondary antibodies were Alexa Fluor conjugates (1:750, 16 hours, Life Technologies). For each retina, the recorded region was identified by the local blood vessel pattern and confirmed by comparing size and position of individual somata in the GCL. Image stacks were acquired on a confocal microscope (Nikon Eclipse C1) equipped with a x60 oil objective (1.4 NA). The degree of immunolabelling of GCL cells was evaluate and rated (from 0, negative, to 4 positive) using z-stacks. Attribution of labelled somata to recorded ones was performed manually using ImageJ (http://imagej.nih.gov/ij) and IGOR Pro.
Baden T., Berens P., Franke K., Rosón M.R., Bethge M, & Euler T. (2016). The functional diversity of retinal ganglion cells in the mouse. Nature, 529(7586), 345-350.
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4

Immunofluorescence Characterization of Endothelial Cell-Tumor Cell Interactions

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Fixation was carried out by adding an equal volume of 2× fixative (PBS with 4% paraformaldehyde and 0.4% glutaraldehyde) to EC monolayers in EGM-2 MV medium 15 min, 1 hr, 3 hrs, and 6 hrs after addition of UM cells. After 15 min at 37°C, cells were permeabilized with 0.1% Triton X-100 in PBS for 5 min, washed with PBS, and blocked with 2% fish gelatin (Sigma-Aldrich) in PBS. Primary and secondary antibodies were diluted in 2% fish gelatin in PBS. Primary antibodies included mouse monoclonal anti-cortactin (Millipore, Billerica, MA), mouse monoclonal anti-E-cadherin (BD Biosciences, Franklin Lakes, NJ), mouse monoclonal anti-N-cadherin (BD Biosciences), mouse monoclonal anti-VCAM (R&D Systems, Minneapolis, MN), rabbit polyclonal anti-S100 (DakoCytomation, Denmark), and rabbit polyclonal anti-VE-cadherin (Cell Signaling, Danvers, MA). F-actin was visualized using Alexa-fluor-conjugated phalloidin (Life Technologies). Secondary antibodies were Alexa-fluor conjugates (Life Technologies), and the mounting agent was ProLong Gold (Life Technologies). Cells were imaged with an inverted microscope (Olympus IX72) using a 40× objective.
Onken M.D., Li J, & Cooper J.A. (2014). Uveal Melanoma Cells Utilize a Novel Route for Transendothelial Migration. PLoS ONE, 9(12), e115472.
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5

Whole-Mount Immunofluorescence of Mouse Trachea

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For Rosa26R-fGFP and wild-type mice, tracheas were fixed overnight in 4% paraformaldehyde at 4°C. Primary antibodies were anti-GFP (chicken, 1:1,000; Abcam, AB13970), anti-KRT5 (rabbit, 1:500; Covance, PRB-160P), anti-acetylated tubulin (mouse, 1:1,000; Sigma, T7451), and anti-PGP9.5 (guinea pig, 1:500; Neuromics, GP14104). Secondary antibodies were Alexa Fluor conjugates (Life Technologies, 1:2,000). Samples were processed to 97% TDE (2′2′-thiodiethanol) for mounting. For mice carrying tetO-H2B-GFP or Rosa26R-confetti, the whole-mount protocol was adapted to enable direct visualization of native fluorescence. Anti-GFP staining was omitted, and samples were mounted in Glycergel (Dako) + 2.5% DABCO.
Watson J.K., Rulands S., Wilkinson A.C., Wuidart A., Ousset M., Van Keymeulen A., Göttgens B., Blanpain C., Simons B.D, & Rawlins E.L. (2015). Clonal Dynamics Reveal Two Distinct Populations of Basal Cells in Slow-Turnover Airway Epithelium. Cell Reports, 12(1), 90-101.
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6

Imaging-Based Quantification of Eukaryotic Stress Response

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GFP-LC3 stable expressing U2OS cells were seeded in 384-well microplates for 24 h. After experimental treatments, cells were fixed with 4% paraformaldehyde for 20 min at room temperature and permeabilized with 0.1% Triton X-100 (v:v in PBS) for 10 min on ice. Thereafter, cells were maintained in 5% bovine serum albumin (BSA, w/v in PBS) for 1 h to block non-specific binding, followed by overnight incubation at 4 °C with phosphoneoepitope-specific eIF2α antibody (ab32157, Abcam, Cambridge, UK). After several washing steps with PBS, cells were incubated in AlexaFluor™ conjugates (Life Technologies) against the primary antibody for 2 h at room temperature. Nuclear staining was achieved by incubation with 10 μg/ml Hoechst 33342 in PBS. Images were acquired and analyzed as described before.
Sauvat A., Chen G., Müller K., Tong M., Aprahamian F., Durand S., Cerrato G., Bezu L., Leduc M., Franz J., Rockenfeller P., Sadoshima J., Madeo F., Kepp O, & Kroemer G. (2018). Trans-Fats Inhibit Autophagy Induced by Saturated Fatty Acids. eBioMedicine, 30, 261-272.
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7

Immunolabeling of Retinal Cell Types

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Following Ca2+ imaging, retinas were mounted onto filter paper (0.8 μm pore size, Millipore) and fixed in 4% paraformaldehyde (in PBS) for 15 min at 4°C. Immunolabelling was performed using antibodies against ChAT (choline-acetyltransferase; goat anti-ChAT, 1:100, AB144P, Millipore), GAD67 (glutamate decarboxylase; mouse anti-GAD67, 1:100, MAB5046, Millipore), SMI-32 (mouse anti-SMI32, 1:100, SMI-32R-100, Covance), and melanopsin (rabbit anti-melanopsin, 1:4000, AB-N38, Advanced Targeting Systems) for 4 days. Secondary antibodies were Alexa Fluor conjugates (1:750, 16 hours, Life Technologies). For each retina, the recorded region was identified by the local blood vessel pattern and confirmed by comparing size and position of individual somata in the GCL. Image stacks were acquired on a confocal microscope (Nikon Eclipse C1) equipped with a x60 oil objective (1.4 NA). The degree of immunolabelling of GCL cells was evaluate and rated (from 0, negative, to 4 positive) using z-stacks. Attribution of labelled somata to recorded ones was performed manually using ImageJ (http://imagej.nih.gov/ij) and IGOR Pro.
Baden T., Berens P., Franke K., Rosón M.R., Bethge M, & Euler T. (2016). The functional diversity of retinal ganglion cells in the mouse. Nature, 529(7586), 345-350.
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8

Antibody Characterization Protocol

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Primary antibodies used were polyclonal rabbit anti-NE81 [12 (link)], monoclonal rat YL1/2 directed against α-tubulin [35 (link)], monoclonal mouse 9E10 directed against myc-tag (EQKLISEEDL) [36 (link)], and monoclonal mouse Act-1 against Dictyostelium actin [37 (link)]. Secondary antibodies involved Alexa Fluor conjugates purchased from Life Technologies (Darmstadt, Germany), Atto conjugates from Atto-Tec GmbH (Siegen, Germany), and enzyme conjugates for Western blotting from Sigma (Deisenhofen, Germany).
Grafe M., Batsios P., Meyer I., Lisin D., Baumann O., Goldberg M.W, & Gräf R. (2019). Supramolecular Structures of the Dictyostelium Lamin NE81. Cells, 8(2), 162.
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9

Immunofluorescence Analysis of Endothelial Cells

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Fixation was carried out by adding 2× fixative (PBS containing 4% paraformaldehyde and 0.4% glutaraldehyde) directly to live cells in tissue culture medium. After 15 min at 37°C, cells were permeabilized with 0.1% Triton X-100 in PBS for 5 min, washed with PBS, and blocked with 3% BSA in PBS. Primary and secondary antibodies were diluted in 3% BSA. Primary antibodies included rabbit anti-VE-cadherin (Cell Signaling, Danvers, MA), mouse anti-human vinculin (Sigma-Aldrich), and mouse anti-human ICAM-1 (R&D Systems, Minneapolis, MN). F-actin was visualized using Alexa-fluor conjugated phalloidin (Life Technologies). Secondary antibodies were Alexa-fluor conjugates (Life Technologies), and the mounting agent was ProLong Gold (Life Technologies). Cells were imaged with an inverted microscope (Olympus IX72) using 40× and 100× objectives.
Onken M.D., Mooren O.L., Mukherjee S., Shahan S.T., Li J, & Cooper J.A. (2015). Endothelial Monolayers and Transendothelial Migration Depend on Mechanical Properties of the Substrate. Cytoskeleton (Hoboken, N.J.), 71(12), 695-706.
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10

Immunofluorescence Staining of TFEB and TFE3

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Cells were fixed with 3.7% paraformaldehyde for 20 min at room temperature, permeabilized with 0.1% Triton on ice, and then blocked with 5% bovine serum albumin (BSA, w/v in PBS) for 1 h, followed by overnight incubation at 4°C with antibodies specific to TFEB (#4240, 1:400, Cell Signaling Technology) or TFE3 (ab93808, 1:200, Abcam). Thereafter, Alexa Fluor™ conjugates (Life Technologies) against the primary antibody were applied for 2 h at room temperature. Cells were then washed and imaged by high‐content microscopy as described above.
Chen G., Xie W., Nah J., Sauvat A., Liu P., Pietrocola F., Sica V., Carmona‐Gutierrez D., Zimmermann A., Pendl T., Tadic J., Bergmann M., Hofer S.J., Domuz L., Lachkar S., Markaki M., Tavernarakis N., Sadoshima J., Madeo F., Kepp O, & Kroemer G. (2019). 3,4‐Dimethoxychalcone induces autophagy through activation of the transcription factors TFE3 and TFEB. EMBO Molecular Medicine, 11(11), e10469.
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