Cells were fixed for 20 min in 4% paraformaldehyde and permeabilized with 0.1% Triton X-100 (in PBS) for 10 min. After three PBS washes, cells were blocked with 2% bovine serum albumin (BSA) for 30 min at ambient temperature. Cells were then incubated with antibodies diluted in 2% BSA at 4°C overnight. After three PBS washes, the cells were incubated with 1 μg/mL Alexa Fluor 488/647-conjugated secondary antibodies (Invitrogen) by shaking at ambient temperature in the dark for 1 h. Cells were washed three times with PBS in the dark and mounted in Prolong Gold Antifade Reagent (Invitrogen). Immunofluorescent staining was observed and analyzed using confocal or fluorescent microscopes (Zeiss) and ZEN software (Zeiss).
Alexa fluor 488 647 conjugated secondary antibodies
Manufactured by Thermo Fisher Scientific
Sourced in United States
Alexa Fluor 488/647 conjugated secondary antibodies are fluorescent-labeled antibodies used as detection reagents in various immunoassays and microscopy techniques. They bind to and detect primary antibodies, enabling visualization and quantification of target proteins or other biomolecules.
Automatically generated - may contain errors
Lab products found in correlation
24 well glass bottom plate, by Cellvis (2 mentions)
Alexa fluor 488 conjugated goat anti rabbit secondary antibody, by Thermo Fisher Scientific (2 mentions)
800 airyscan microscope, by Zeiss (2 mentions)
Image express micro confocal high content imaging system, by Molecular Devices (2 mentions)
Rabbit anti phospho upf1 antibody, by Merck Group (2 mentions)
Zen software, by Zeiss (1 mentions)
Prolong gold antifade reagent, by Thermo Fisher Scientific (1 mentions)
Ab72965, by Abcam (1 mentions)
Ap06779, by OriGene (1 mentions)
Fluoview fv1000, by Olympus (1 mentions)
5 protocols using alexa fluor 488 647 conjugated secondary antibodies
1
Immunofluorescent Staining of Fixed Cells
2
Chemerin and CMKLR1 Localization Comparison
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The localizations and expression of chemerin and CMKLR1 were compared using immunofluorescence (IF) staining. Tissues were stained for 2 h at room temperature in the presence of anti-chemerin (1 : 200, ab72965, Abcam, USA) and anti-CMKLR1 (1 : 200, AP06779, Origene, USA) antibodies, followed by staining for 1 h at room temperature with appropriate Alexa Fluor 488/647-conjugated secondary antibodies (Invitrogen, Carlsbad, CA, USA). Tissue sections were stained with DAPI for 5 min before imaging.
3
Immunofluorescence Staining of HEK293T and iPSNs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
HEK293T cells were fixed and stained as previously described (Coyne et al., 2020 (link)) using Rabbit anti-phospho-UPF1 antibody (Millipore Sigma) at a 1:250 dilution and AlexaFluor 488 conjugated Goat anti-Rabbit secondary antibody (Thermo Fisher Scientific) at a 1:1000 dilution in a 24-well glass-bottom plate (Cellvis). 9 sites in each well were imaged using a 20x objective on an ImageExpress Micro Confocal High-Content Imaging System (Molecular Devices).
iPSNs were fixed with 4% (v/v) para-formaldehyde in PBS for 10 min, permeabilized in 0.2% (v/v) Triton X-100 for 10 min, blocked in 1% bovine serum albumin and 2% goat serum for 1h, incubated with primary antibodies overnight at 4°C, washed with PBS, and finally incubated with Alexa Fluor 488/647 conjugated secondary antibodies (ThermoFisher Scientific). Cells were imaged with a Zeiss 800 Airyscan microscope.
iPSNs were fixed with 4% (v/v) para-formaldehyde in PBS for 10 min, permeabilized in 0.2% (v/v) Triton X-100 for 10 min, blocked in 1% bovine serum albumin and 2% goat serum for 1h, incubated with primary antibodies overnight at 4°C, washed with PBS, and finally incubated with Alexa Fluor 488/647 conjugated secondary antibodies (ThermoFisher Scientific). Cells were imaged with a Zeiss 800 Airyscan microscope.
4
Immunofluorescent Staining of Liver Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Immunofluorescent staining of liver tissues was performed as we reported.50 Briefly, liver tissues were fixed with 10% neutral buffered formalin and cut into sections of 5-um thickness. After the non-specific binding sites were blocked with 10% donkey serum for 1 hour at room temperature, the sections were incubated with anti-α-SMA antibody, anti-CD68 antibody, anti-desmin, anti-(GFAP), and anti-collagen type I (for details of primary antibodies, see Table 2 ) at 4 °C overnight, washed extensively in phosphate buffered saline, with Alexa Fluor 488/647 conjugated secondary antibodies (1:500, ThermoFisher, A-21206/A-31573) for 1 hour. Sections were then counterstained with 4, 6-diamidino-2-phenylindole (DAPI), washed, and mounted for observation under a scanning confocal microscope (Olympus, Fluoview FV1000).
5
Immunofluorescence Staining of HEK293T and iPSNs
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!