To determine the co-localization between α-syn or neurosin and cellular markers, double-labeling experiments were performed as previously described [25 (link)]. Vibratome sections were immunolabeled with antibodies against S100 (astroglia), Iba1 (microglia), Olig-2 (oligodendrocytes), p25 (oligodendrocytes) or NeuN (neurons), and the immunoreactive structures were detected with FITC-tagged secondary antibody (Vector Laboratories, 1:75) and the V5 tagged neurosin with the Tyramide Signal Amplification™-Direct system (NEN Life Sciences), while α-syn was detected with an antibody specific for human α-syn (SYN211, Sigma) and a FITC-tagged secondary antibody (Vector Laboratories, 1:75). Cell nuclei were stained using ProLong® Gold Antifade with DAPI (4',6-diamidino-2-phenylindole) (Molecular Probes). Sections were imaged with a Zeiss 63X objective on an Axiovert 35 microscope (Zeiss) with an attached MRC1024 laser scanning confocal microscope (BioRad) with an optical image of 1 μm thick with fluorescent signals in co-registry [60 (link)].
Fitc tagged secondary antibody
Manufactured by Vector Laboratories
The FITC-tagged secondary antibody is a fluorescent-labeled detection reagent. It is designed to recognize and bind to the Fc region of primary antibodies, allowing for sensitive visualization and detection of target proteins or molecules in various immunoassays and imaging applications.
Automatically generated - may contain errors
Lab products found in correlation
Axiovert 35 microscope, by Zeiss (3 mentions)
Mrc 1024 laser scanning confocal microscope, by Bio-Rad (3 mentions)
Syn211, by Merck Group (2 mentions)
Prolong gold antifade with 4 6 diamidino 2 phenylindole dapi, by Thermo Fisher Scientific (1 mentions)
Tcs spe confocal system, by Leica (1 mentions)
Dmi4000 b inverted fluorescent microscope, by Leica (1 mentions)
4 protocols using fitc tagged secondary antibody
1
Dual Immunolabeling for Co-localization Analysis
2
Colocalization of TH and BrdU in Grafted NSCs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To determine the colocalization between TH and BrdU in grafted NSCs, double-labeling experiments were performed. Briefly, vibratome sections were immunolabeled with antibodies against TH (1:250; Millipore) and BrdU (1:500; Millipore). The TH was detected with an FITC-tagged secondary antibody (1:75; Vector Laboratories) while BrdU with the Tyramide Signal Amplification™-Direct system (1:100; NEN Life Sciences). Sections were imaged with a Zeiss 63× objective on an Axiovert 35 microscope (Zeiss) with an attached MRC 1024 laser scanning confocal microscope (BioRad).
3
Colocalization of α-Synuclein with Neuronal and Glial Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To determine the colocalization between α-syn and neuronal (NeuN) and glial markers (Iba1, S100, p25), double-labeling experiments were performed as previously described [28 (link)]. Vibratome sections were immunolabeled with antibodies against S100 (astroglia, Sigma, 1:250), Iba1 (microglia), p25 (oligodendrocytes) or NeuN (neurons), and the immunoreactive structures were detected with the Tyramide Signal Amplification™-Direct system (1:100, NEN Life Sciences), while α-syn was detected with an antibody specific for human α-syn (SYN211, Sigma, 1:250) and a FITC-tagged secondary antibody (Vector Laboratories, 1:75). Cell nuclei were stained using ProLong® Gold Antifade Mountant with DAPI (4′,6-diamidino-2-phenylindole) (Molecular Probes). Sections were imaged with a Zeiss 63X objective on an Axiovert 35 microscope (Zeiss) with an attached MRC1024 laser scanning confocal microscope (BioRad) [56 (link)].
4
Quantifying α-Synuclein Cross-Seeding
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For the double labeling studies, 40 μm sections were immunolabeled with the Syn1 antibody and either MAP2 (Clone 2B, Millipore, Cat. No: MAB378), Rab5 (15/Rab5, BD Biosciences, Cat. No: 610,281), GFAP (Millipore, Cat. No: MAB3402) or a mouse specific α-syn antibody (a generous gift from V.M-Y. Lee). Human α-syn immunoreactivity was detected with a FITC-tagged secondary antibody (Vector), while the other antibodies were visualized with the Tyramide Signal Amplification™-Direct (Red) system (Perkin Elmer). All sections were processed simultaneously under the same conditions, and experiments were performed in duplicate in order to assess the reproducibility of results. All fluorescent imaging studies were done on a DMI 4000B inverted fluorescent microscope (Leica, Germany) with an attached TCS SPE confocal system (Leica), using a Leica 63X (N.A. 1.3) objective. A total of four optically paired micrographs were analyzed with ImageJ colocalization color map plugin to determine the percent of human α-syn positive clusters within Rab5, MAP2, and GFAP expressing cells. Efficacy of of α-syn cross-seeding was reported as percent mouse α-syn clusters detected within human α-syn positive cells.
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!