Medaka larvae at 10 dpf were fixed in 4% paraformaldehyde in 0.85X PBS at 4 °C overnight. After fixation, the specimens were embedded in paraffin and sectioned serially at 5 μm. The deparaffinized sections were treated with anti-GFP antibody (1:500, Invitrogen, A11122) at room temperature for 3 h. After several washes with PBS, sections were incubated with Alexa 488-conjugated anti-rabbit IgG secondary antibody (1:500, Invitrogen, A21206) for 1 h. The sections were counterstained with DAPI.
Alexa 488 conjugated anti rabbit igg secondary antibody
Manufactured by Thermo Fisher Scientific
Sourced in United States
Alexa Fluor® 488-conjugated anti-rabbit IgG secondary antibody is a fluorescent-labeled secondary antibody used to detect and visualize rabbit primary antibodies in various immunoassay techniques, such as immunohistochemistry, immunocytochemistry, and Western blotting.
Automatically generated - may contain errors
Lab products found in correlation
Anti gfp antibody, by Thermo Fisher Scientific (1 mentions)
Tsa cy3 system, by PerkinElmer (1 mentions)
Axio imager z2, by Zeiss (1 mentions)
Peroxidase conjugated secondary antibody, by Santa Cruz Biotechnology (1 mentions)
Draq5, by Thermo Fisher Scientific (1 mentions)
Hrp labeled secondary antibody, by Vector Laboratories (1 mentions)
3 3 diaminobenzidine (dab), by Vector Laboratories (1 mentions)
Anti lc3 2, by Cell Signaling Technology (1 mentions)
Supersignal enhanced chemiluminescence ecl reagent, by Cytiva (1 mentions)
Lysotracker red dnd 99, by Thermo Fisher Scientific (1 mentions)
Pvdf membrane, by Merck Group (1 mentions)
Fsx100 fluorescence microscope, by Olympus (1 mentions)
Anti α tubulin, by Merck Group (1 mentions)
Anti γh2ax, by Cell Signaling Technology (1 mentions)
Sm2020 r, by Leica (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Biotium (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Keyence (1 mentions)
Bz x710, by Keyence (1 mentions)
6 protocols using alexa 488 conjugated anti rabbit igg secondary antibody
1
GFP Immunohistochemistry in Medaka Larvae
2
Quantitative Phospho-S6 Ribosomal Protein Imaging
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System; PerkinElmer) for 1 hat RT. The cells were incubated with anti-digoxigenin-POD (1 : 200 in TNB, Roche) and rabbit anti-Phospho-S6 Ribosomal Protein antibody (1 : 400 in TNB, #4858S, Cell Signaling) in a humidified chamber overnight in 4°C, next they were washed with PBS-T, and incubated with Alexa 488-conjugated anti-rabbit IgG secondary antibody (1 : 1000; Invitrogen). The hybridization signal was amplified with TSA Cy3 System (Perkin Elmer). The imaging was performed using confocal microscope LSM 700, Axio Imager Z2 (Zeiss) in a stack of 0.4 μm thick Z section, using a 40× objective (oil immersion; 1.3 NA) with 1.2 zoom factor. The obtained stack was "flattened" into a single image using a maximum intensity projection. Quantification of fluorescence signal intensity was performed in ImageJ software. Signal intensity was averaged on the cell area.
3
Autophagy-Related Protein Detection
4
Immunofluorescence Staining of Tubulin and γH2AX
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Cells were cultured on cover glasses in well plates. The prepared cover glasses were fixed in 4% paraformaldehyde/phosphate-buffered saline (PBS) at 25 °C for 15 min and washed three times with PBS. The cells were permeabilized and blocked with 0.3% Triton X-100 (Agilent Technologies, Santa Clara, CA, USA) and blocked with 5% FBS at room temperature for 60 min. The cells were then washed three times with PBS and incubated with anti-α-tubulin (1:1000 #T6199 Sigma Aldrich, St Luis, Missouri, USA) and anti-γH2AX (1:500 #2578 Cell signaling Technology, Danvers, MA, USA) at 4 °C overnight. After extensive washing with PBS, the samples were incubated with Alexa-594-conjugated goat anti-mouse IgG secondary antibody (1:500 #A11001 Life technologies,) and Alexa-488-conjugated anti-rabbit IgG secondary antibody (1:500 #A11072 Life technologies) at 25 °C for 60 min. The cover glasses were washed three times with PBS, mounted in in VECRASHELD Mounting medium for fluorescence (Vector Laboratories, Inc Burlingame, CA), and sealed with nail polish. Images were acquired using OLYMPUS FSX 100 fluorescence microscope (OLYMPUS, Tokyo, Japan).
5
Immunohistochemistry of Neuronal Cell Types
6
Quantifying Neurite Outgrowth from Thawed Neurospheres
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Neurospheres frozen on day 27 were thawed, seeded in 24-well plates coated with iMatrix511 for three days, and fixed with 4% paraformaldehyde. The neurospheres were stained with an anti-tubulin beta III (TUBB3) antibody (BioLegend, Inc., San Diego, CA, USA) and an Alexa 488-conjugated anti-rabbit IgG secondary antibody (Thermo Fisher Scientific), followed by counterstaining of the nucleus with 4’,6-diamidino-2-phenylindole (DAPI) (Biotium Inc., Fremont, USA). A fluorescence microscope (BZ-X710; KEYENCE CORPORATION, Osaka, Japan) was used for visualization, and the area covered by neurites was defined as the TUBB3-positive area minus the DAPI-positive area using the BZ-H3M measurement module (KEYENCE CORPORATION).
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