Prior to injection, larval zebrafish were anesthetized with 0.01% tricaine, and immobilized in 1.5% low melting‐point agarose. 1–2 nL of rVSV‐Venus(VSV‐G), 1 × 106 or 1 × 108 ffu/mL, rVSV‐GFP(RABV‐G), 1 × 108 ffu/mL, or rVSVΔG‐GFP(RABV‐G), 1 × 108, or 1 × 109 ffu/mL was injected into the vitreal cavity of one eye. Infected animals were fixed at 24, 48, or 72 hpi and processed by whole‐mount immunohistochemistry. Samples were incubated with rabbit anti‐GFP to detect Venus or GFP (MBL International, Woburn, MA; 598, RRID:AB_591819), mouse anti‐HU (HuC/D) (Life Technologies, Bethesda, MD; RRID:AB_591819), and mouse anti‐HNK1 (Developmental Studies Hybridoma Bank, Iowa City, IA; RRID:AB_531908) followed by secondary antibodies conjugated with Alexa dyes (Life Technologies, 1:500). Stained samples were mounted in 1.5% low‐melting‐point agarose and imaged with confocal microscopy. 3D rendering of zebrafish larvae was generated with FluidVis (Fluidity Software, Somerville, MA), followed by manual annotation in ImageJ (NIH, Bethesda, MD; RRID:nif‐0000‐30467) and Photoshop.
Alexa dye
Manufactured by Thermo Fisher Scientific
Sourced in United States
Alexa dyes are a series of fluorescent dyes developed by Thermo Fisher Scientific. They are designed to be used as labeling reagents for various biological applications, such as flow cytometry, microscopy, and immunoassays. Alexa dyes offer a range of excitation and emission wavelengths, allowing for flexibility in experimental design. Their photostability and brightness make them suitable for a variety of research applications.
Automatically generated - may contain errors
Lab products found in correlation
Dabco mowiol, by Thermo Fisher Scientific (6 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (4 mentions)
Formaldehyde, by Electron Microscopy Sciences (4 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (3 mentions)
Polyclonal chicken anti map2, by Abcam (3 mentions)
Ab97959, by Abcam (3 mentions)
Operetta, by PerkinElmer (3 mentions)
Mouse anti hnk1, by Developmental Studies Hybridoma Bank (2 mentions)
Mouse anti hu huc d, by Thermo Fisher Scientific (2 mentions)
Lsm 700, by Zeiss (2 mentions)
Vectastain elite abc kit, by Vector Laboratories (2 mentions)
Anti gfp, by Abcam (2 mentions)
Lsm 710, by Zeiss (2 mentions)
Lsm 880 confocal microscope, by Zeiss (2 mentions)
G3893, by Merck Group (2 mentions)
Lsm 510 microscope, by Zeiss (2 mentions)
Polyclonal guinea pig vglut1, by Merck Group (2 mentions)
Mab1259, by Abcam (2 mentions)
Lsm 510, by Zeiss (2 mentions)
Lsm 510 meta, by Zeiss (2 mentions)
49 protocols using alexa dye
1
Tracing Virus Spread in Zebrafish Retina
2
Comprehensive Antibody Labeling Protocol
3
Autofluorescence Reduction in Tissue Sections
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Fresh frozen sections were post-fixed using 4% paraformaldehyde (PFA) in phosphate buffered saline (PBS). After a brief wash in PBS, sections were stained by Sudan Black B for elimination of the autofluorescence due to lipofuscin [12 (link)]. Following immersion in 10% goat serum in PBS and 0.1% Tween20 (PBS-T) for 60 min, the sections were incubated with primary antibodies diluted in 1% BSA-PBS-T for 18 h at room temperature. Sections were then rinsed with PBS-T, and bound antibodies were visualized with secondary antibodies conjugated with Alexa dyes (Life Technology). The specimens were visualized by confocal-laser-scanning-microscope (LSM 700; Carl Zeiss Inc.). Whenever necessary, bound antibodies were labeled by biotinylated anti-mouse or anti-rabbit IgG antibodies (Vector Laboratories, Inc., Burlingame, CA), followed by avidin and biotinylated HRP complex (Vectastain Elite ABC kit; Vector Laboratories, Inc.). Bound HRP was developed with 3,3-diaminobenzidine (DAB) in the presence of hydrogen peroxide.
4
Viral Tracing in Larval Zebrafish
5
Photoconversion of Fluorescent Dyes
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For photoconversion of dyes alone, 3 μL of dye was mixed with 1 μL of 3 μm polystyrene beads (10−4% v/v) and dispensed on a glass slide, covered with a #1.5 coverslip and placed under the objective lens. The final concentration of SYTO dyes (Life Technologies) used (following dilution) was 3.75 mM. Alexa dyes (Life Technologies) were used at 7.5 g/L. Cy 5 (Lumiprobe) was used at 0.1 g/L. All other cyanine dyes (Lumiprobe) were used at 10 g/L. To quantify green enhancement from photoconversion, the sample was exposed to the femtosecond laser delivering 810 nm light at a power of 80 mW for up to 30 s. The bleaching of the initial red fluorescence (ΔRi) and signal enhancement in the green or blue channels by photoconversion (ΔF) were quantified to calculate conversion brightness (ΔF) and conversion yield (ΔF/ ΔRi).
6
Immunofluorescence Assay for LC3-II
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MDA-MB-231 cells were plated and transfected with siRNA on chamber slides (Thermo Fisher Scientific, Rockville, MD). Following drug treatment, cells were fixed with 4% paraformaldehyde in PBS and subsequently permeabilized with 100 μg/mL digitonin in PBS. Cell were treated with 1% BSA and incubated with anti-LC3-II (MBL, Woburn, MA), followed by secondary antibody conjugated with Alexa dyes (Life Technologies, Carlsbad, CA). Slides were mounted with cover-slips with DAPI-containing mounting solution (Vector Laboratories, Burlingame, CA). Digital images were acquired with a Zeiss LSM 700 confocal system using a 63X oil-immersion objective.
7
Immunostaining of Retina Sections and Flatmounts
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Preparation and immunostaining of retina sections and whole retina flatmounts was performed as described previously15 (link)40 (link). Primary antibodies were incubated for 24–72 hours at 4 °C. Secondary antibodies were incubated 1:200 for 2 hours at 22 °C. All secondary antibodies were raised in donkey and conjugated with Alexa dyes (Life Technologies). A summary of primary and secondary antibodies is shown in Table 1 . For retinal sections, all antibodies were diluted in PBS with 2.5% donkey serum and 0.2% Triton-X. All wash steps were performed using PBS with 0.05% Tween-20. For staining of retina flatmounts levels of Triton-X were increased to 1%. Samples were mounted in Prolong Gold anti-fade media containing DAPI (Life Technologies).
8
Embryo Immunostaining and Imaging
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Embryos were fixed by 4% formaldehyde (37% for microtubules staining) and stained according to standard procedures (22 (link)). F-actin was detected with labeled phalloidin (Invitrogen, Carlsbad, CA) and DAPI (0.2 μg/mL). The following antibodies were employed: monoclonal anti-α-Tubulin (Sigma-Aldrich, St. Louis, MO) and goat IgGs coupled with Alexa dyes (at final 4 μg/mL, Invitrogen).
9
Immunofluorescence Staining of Caveolin-1 and Fibrillin-2
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were fixed at -20°C with 70% MetOH 30% Acetone 5min, blocked 30min with 10% FBS in PBS and stained with anti-Caveolin1 (1:5000) and anti-Fbn2 (1:150) overnight at 4°C. After several washes with PBS, cells were incubated 1hr at room temperature with the appropriate secondary antibodies conjugated with Alexa dyes (Invitrogen) diluted 1:500, washed several times with PBS and mounted in ProLong Gold antifade reagent (Molecular Probes).
10
Immunostaining Embryonic Tissue Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Embryos were fixed, stained, and mounted as previously described 42 . Antibodies against the following antigens were used: Dlg (mouse, 0.4 μg/ml) 43 , Arm (mouse M7A1, 0.4 μg/ml) 44 , and Slam (rabbit, 1:5,000) 45 . Secondary antibodies were labeled with Alexa dyes (Invitrogen, 0.4 μg/ml). GFP booster labeled with ATTO488 (ChromoTek, 1:500) was used for E‐Cad‐GFP.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!