Alexa fluor 488 conjugate

Manufactured by Jackson ImmunoResearch

Alexa Fluor 488 is a fluorescent dye that can be conjugated to antibodies, proteins, or other biomolecules. It has an excitation maximum of 495 nm and an emission maximum of 519 nm, making it suitable for detection in flow cytometry, fluorescence microscopy, and other fluorescence-based applications.

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Lab products found in correlation

F4680, by Merck Group (2 mentions) Cyanine3 (cy3), by Jackson ImmunoResearch (2 mentions) Goat anti rabbit igg, by Jackson ImmunoResearch (1 mentions) Goat anti mouse igg, by Jackson ImmunoResearch (1 mentions) Goat anti rat igg, by Jackson ImmunoResearch (1 mentions) Rabbit anti phospho jnk thr183 tyr185, by Cell Signaling Technology (1 mentions) Rabbit anti dcp 1 asp216, by Cell Signaling Technology (1 mentions) Rabbit anti gfp tag polyclonal, by Thermo Fisher Scientific (1 mentions) Alexa fluor 555 conjugate, by Jackson ImmunoResearch (1 mentions) Prolong gold antifade reagent, by Thermo Fisher Scientific (1 mentions) Neurotrace 530 615 red fluorescent nissl stain, by Thermo Fisher Scientific (1 mentions) Fluoview fv3000 confocal laser scanning microscope, by Olympus (1 mentions) Nb120 16007, by Novus Biologicals (1 mentions) Sc 514592, by Santa Cruz Biotechnology (1 mentions)

Spelling variants of this product

Alexa Fluor 488 (424 citations) Alexa 488 (166 citations) Donkey anti-rabbit IgG Alexa Fluor 488 conjugate (2 citations)

6 protocols using alexa fluor 488 conjugate

Drosophila Embryo Immunostaining Protocol

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Immunostaining of flat-prepped stage 16 Drosophila embryos was performed using the following primary antibodies: mouse anti-Fas 2 (1:100, clone 1D4, DHSB), mouse anti-axons CNS (1:100, BP 102, DHSB), mouse anti-Futsch (1:1000, 22c10, DHSB), rabbit anti-phospho-JNK (Thr¹⁸³ / Tyr¹⁸⁵) (1:100, Cell Signaling #9251), rabbit anti-Dcp-1 (Asp216) (1:100, Cell Signaling #9578), rabbit anti-GFP tag polyclonal (1:600, Thermo Fisher Scientific), rat anti-Elav (1:1000, clone 7E8A10, DHSB), mouse anti-Repo (1:100, clone 8D12 DHSB), mouse anti-Even-skipped (1:100, clone 3C10, DHSB), mouse anti-Engrailed (1:100, clone 4D9, DHSB) and TRITC-conjugated goat anti-HRP (1:200, Jackson ImmunoResearch #123-025-021).
The secondary antibodies used for detection were: Goat anti-Rabbit IgG (H + L), Alexa Fluor 488 conjugate (A-11008), Goat anti-Rabbit IgG (H + L) Alexa Fluor 555 conjugate (A-21428), Goat anti-Mouse IgG (H + L) Alexa Fluor 488 conjugate (A-11001), Goat anti-Mouse IgG (H + L) Alexa Fluor 555 conjugate (A-21422) and Goat anti-Rat IgG (H + L) Alexa Fluor 555 conjugate (A-21434). All secondary antibodies were used in a dilution of 1:600 and were from Invitrogen.

Karkali K., Saunders T.E., Panayotou G, & Martín-Blanco E. (2023). JNK signaling in pioneer neurons organizes ventral nerve cord architecture in Drosophila embryos. Nature Communications, 14, 675.

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Immunofluorescence Staining of Brain Slices

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Free‐floating slices were rinsed in PBS then permeabilized and blocked with PBS‐BT (50 mM Tris–HCl, 150 mM NaCl, 3% bovine serum albumin (BSA), 0.1% Triton X‐100, pH 7.4) blocking solution for 1 h. Afterward, the sections were incubated with primary antibodies (see key resource table) in a 2% normal donkey serum (NDS) and 0.3% Triton X‐100 PBS solution on a shaker overnight at 4°C. The next day, sections were rinsed and incubated with corresponding secondary antibodies directly conjugated with fluorophores (1:500, Cy3, Cy5, and Alexa Fluor 488 conjugate from Jackson ImmunoResearch) for 2 h at room temperature. Finally, slices were rinsed in PBS and mounted (Sigma‐Aldrich, F4680). For DAT and netrin‐1 immunostaining, antigen retrieval was performed in citrate buffer (pH 9) at 80°C.

Jasmin M., Ahn E.H., Voutilainen M.H., Fombonne J., Guix C., Viljakainen T., Kang S.S., Yu L., Saarma M., Mehlen P, & Ye K. (2020). Netrin‐1 and its receptor DCC modulate survival and death of dopamine neurons and Parkinson’s disease features. The EMBO Journal, 40(3), e105537.

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Enhancing GFP Signal in cFos-eGFP Brains

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To amplify and increase longevity of the GFP signal in cFos tagged cells of the cFos-eGFP brains, one series of sections from each region of interest (BLA, Auditory Cortex (AU), BNST, Nucleus Accumbens Dorsal Core (AcDC), Nucleus Accumbens Medial Core (AcC), and Nucleus Accumbens Shell (AcS); each section spaced ~240mm apart) were subjected to GFP immunohistochemistry. Free-floating sections were washed 3×15 min in 1x PBS and incubated with blocking buffer (0.3% TritonX-100 PBS with 3% normal donkey serum) for 1.5 hours. They were then incubated in chicken-anti-GFP igY (GFP-1020, Aveslabs) diluted 1:1000 in blocking buffer for ~12 hours. The sections were washed 3×15 min in 1x PBS and incubated in donkey anti-chicken Alexa Fluor488 conjugate (703–545–155, Jackson ImmunoResearch) for 8 hours, followed by a 3×15 min wash in 1x PBS. For better visualization of the regions of interest, a fluorescent NISSL stain step was added following cFos staining. Sections were incubated in NeuroTrace 530/615 red fluorescent NISSL stain (Invitrogen, N21482) diluted 1:200 in 1x PBS for one hour and then washed 3×15 min in 1x PBS. All aforementioned procedures were done at room temperature. Six slices per region of interest were mounted and coverslipped with ProLong Gold antifade reagent (Invitrogen, P36930) for later microscopy.

Demaestri C., Brenhouse H.C, & Honeycutt J.A. (2018). 22kHz and 55kHz ultrasonic vocalizations differentially influence neural and behavioral outcomes: Implications for modeling anxiety via auditory stimuli in the rat. Behavioural brain research, 360, 134-145.

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Immunohistochemistry and Immunofluorescence of Murine Brain

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Mice were anesthetized and then transcardially perfused with cold PBS and 4% paraformaldehyde (PFA). The brains were stored for 24 h in 4% PFA at 4°C and then embedded in paraffin. Serial 5-μm-thick sections from all animal groups were processed in parallel for immunohistochemistry and immunofluorescence. Free-floating slices were rinsed in PBS then permeabilized and blocked with PBS-BT [50 mM Tris-HCL, 150 mM NaCl, 3% bovine serum albumin (BSA), 0.1% Triton-X100, pH 7.4] blocking solution for 1 h. Afterward, the sections were incubated with primary antibodies (see Supplementary Table 1) in a 2% normal donkey serum (NDS) and 0.3% Triton X-100 PBS solution on a shaker overnight at 4°C. The next day, sections were rinsed and incubated with corresponding secondary antibodies directly conjugated with fluorophores (1:3000, Cy3, Cy5, and Alexa Fluor 488 conjugate from Jackson ImmunoResearch) for 2 h at room temperature. Finally, slices were rinsed in PBS and mounted (Sigma Aldrich, F4680).

Lee Y.J., Jeong Y.J., Kang E.J., Kang B.S., Lee S.H., Kim Y.J., Kang S.S., Suh S.W, & Ahn E.H. (2023). GAP-43 closely interacts with BDNF in hippocampal neurons and is associated with Alzheimer's disease progression. Frontiers in Molecular Neuroscience, 16, 1150399.

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Quantification of Enteroendocrine Cell Subsets

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To reveal EE and EC phenotypes, monolayers in 24-well inserts were fixed with 4% paraformaldehyde for 15 min, permeabilized with 0.5% Triton-X 100 for 20 min, then blocked in 1% bovine serum albumin (BSA) for 1 h. The samples were subsequently co-stained with chromogranin A (Abcam, ab15160, rabbit, 1:1000) and 5HT (Novus Bio, NB120–16007, mouse 1:200) antibodies in 1% BSA for 16 h at 4°C, followed by staining with secondary antibodies (1:500) of goat anti-mouse (Alexa Fluor 488 conjugate, Jackson Immunoresearch, 115-545-003) and goat anti-rabbit (Alexa Fluor 647 conjugate, Jackson Immunoresearch, 111-605-003) for 1 h. Finally, nuclei were stained with Hoechst 33342 (2 μg/mL) for 15 min. The inserts were placed on a coverglass and the monolayers were imaged with an Olympus FluoView FV3000 confocal laser scanning microscope. The entire surface of inserts (surface area = 33 mm²) was scanned and imaged. The number of EE (ChgA⁺) and EC (5HT⁺) cells were counted using Image J software. Three inserts were used for each condition to obtain the average cell density.

Wang Y., Sims C.E, & Allbritton N.L. (2020). Enterochromaffin Cell–Enriched Monolayer Platform for Assaying Serotonin Release from Human Primary Intestinal Cells. Analytical chemistry, 92(18), 12330-12337.

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Immunodetection Assay in 24-well Plate

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For immunodetection assays in a 24-well plate, the monolayers were fixed with 4% paraformaldehyde for 15 min, followed by permeabilization with 0.5% Triton-X 100 for 20 min and blocked with 1% bovine serum albumin (BSA) at 25 °C for 1 h. The antibodies used for immunodetection were against chromogranin A (ChgA) (Abcam, ab15160, rabbit, 1: 1000), peptide YY (PYY) (Proteintech, 24294-1AP, rabbit 1:200), GLP-1 (Santa Cruz, sc-514592, mouse 1:200). All antibodies were diluted in IF wash (0.1% BSA, 7.7 mM sodium azide, 0.2 % Triton-X, and 0.05% Tween-20 in 1x PBS) and were incubated with the sample at 4 °C for 18 h, followed by staining with secondary antibodies (1: 500) of goat anti-mouse (Alexa Fluor 488-conjugate, Jackson Immunoresearch, 115-545-003) and goat anti-rabbit (Alexa Fluor 594-conjugate, Jackson Immunoresearch, 111-585-003) at 25 °C for 1 h. Finally, cellular DNA was stained with Hoechst 33342 (2 μg/mL) for 15 min.

Villegas-Novoa C., Wang Y., Sims C.E, & Allbritton N.L. (2022). Development of a Primary Human Intestinal Epithelium Enriched in L-cells For Assay of GLP-1 Secretion. Analytical chemistry, 94(27), 9648-9655.

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