Brain sections (20-μm cryostat brain sections) were stained with Fluoro-Jade C to detect neuronal death as described previously.18 (link) Briefly, brain slides were incubated in 100% ethanol (3 min), 70% ethanol (1 min), distilled water (1 min), and then a solution containing 0.0004%, Fluoro-Jade C (Merck Millipore, Darmstadt, Germany), and 0.1% acetic acid for 30 min. Sections were washed three times in distilled water, then dried overnight before being immersed in xylene (3 × 2 min) and mounted with DPX neutral mounting medium (Sigma-Aldrich, St. Louis, MO). Three coronal brain sections (200 μm between slices, −1.2 and −2.2 mm from bregma) from each mouse were imaged with a fluorescence microscope (BX63; Olympus, Tokyo, Japan). Fluoro-Jade C–positive cells were counted using the CellSens Count and Measure Solution module (Olympus) in the perilesional region of the ipsilateral hemisphere by an investigator blind to the treatment.
Fluoro jade c
Manufactured by Merck Group
Sourced in United States
Fluoro-Jade C is a fluorescent stain used to detect neuronal degeneration in tissue sections. It is a sensitive marker for the identification of degenerating neurons. Fluoro-Jade C binds to the cytoplasm of degenerating neurons, allowing for their visualization under a fluorescent microscope.
Automatically generated - may contain errors
Lab products found in correlation
Dibutylphthalate polystyrene xylene (dpx), by Merck Group (4 mentions)
Axio observer z1, by Zeiss (3 mentions)
Lsm 780 confocal microscope, by Zeiss (3 mentions)
Fluoro jade c, by Olympus (2 mentions)
Superfrost plus microscope slide, by Thermo Fisher Scientific (2 mentions)
Dpx mounting medium, by Merck Group (2 mentions)
Fluoro jade, by Merck Group (2 mentions)
Dpx mounting media, by Merck Group (2 mentions)
Entellan, by Merck Group (2 mentions)
Lsm 510, by Zeiss (2 mentions)
Glial fibrillary acidic protein (gfap), by Merck Group (2 mentions)
Fura 2 am, by Thermo Fisher Scientific (2 mentions)
Dp71 digital camera, by Olympus (1 mentions)
Bx51 microscope, by Olympus (1 mentions)
Fluoromount g, by Merck Group (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Santa Cruz Biotechnology (1 mentions)
Potassium permanganate, by Merck Group (1 mentions)
Tcs sp8 laser scanning microscope, by Leica camera (1 mentions)
Dpx mountant, by Merck Group (1 mentions)
Eosin alcohol solution, by Fujifilm (1 mentions)
48 protocols using fluoro jade c
1
Fluoro-Jade C Staining for Neuronal Death
2
Fluoro-Jade C Labeling of Spinal Cord
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Paraffin-embedded slides were deparaffinized, rehydrated, and incubated at room temperature in 0.06% KMnO4 solution for 10 min. Slides were incubated in 0.0001% Fluoro-Jade C (EMD Millipore) dissolved in 0.1% acetic acid for 10 min with gentle shaking. Slides were washed, counterstained with DAPI, and dried on a slide warmer at 50 °C for 5 min. Slides were cleared in xylene for 1 min and then mounted using Cytoseal 60 mounting medium (Fisher Scientific). Slides were imaged using an Olympus BX51 microscope mounted with a DP71 Olympus digital camera. One slide was stained per mouse (each slide contained 2–4 lumbar spinal cord coronal sections). The total number of Fluoro-Jade C-labeled puncta was manually counted for all the spinal cord sections of any one sample and then averaged for each cohort.
3
FluoroJade C Staining of Degenerating Neurons
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The brain tissue preparation for FluoroJade C (FJC; EMD Millipore) was treated exactly as that of the CV brain tissue. Slides were first immersed in 1% sodium hydroxide in 80% ethanol, 70% ethanol and finally in ddH2O before being incubated in a 0.06% potassium permanganate (Sigma-Aldrich) solution. This was followed by an incubation in a 0.0001% FJC solution dissolved in 0.1% aqueous acetic acid and combined with 0.0001% DAPI (Santa Cruz Biotechnology), and slides were once again rinsed with ddH2O and left to air dry. Slides were then immersed in xylene and mounted with FluoroMountG (Sigma-Aldrich) (Ehara and Ueda, 2009 (link)). Imaging was completed with the Zeiss AxioObserver Z1 inverted epifluorescence microscope using GFP (488/509 nm) and DAPI (359/461 nm) filters. Analysis of the total number of degenerating neurons was performed using IMARIS 9.2 (Bitplane). IMARIS 9.2 was set to detect and count all green (representing degenerating neurons) and blue (representing nuclear DNA) spots on each image and then calculate colocalization. The cells having been tagged by both DAPI and FJC were counted as FJC positive neurons.
4
Fluoro-Jade C Staining of Hippocampal Degeneration
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Hippocampal cryo-sections were immersed in a solution of 5% (w/v) NaOH in 80% (v/v) ethanol for 5 min, then sequentially transferred to 70% (v/v) ethanol for 2 min, distilled water for 2 min, 0.06% (w/v) potassium permanganate for 10 min, and distilled water for 2 min at RT. Slices were then incubated for 20 min with 0.0002% (w/v) Fluoro-Jade C (Merck Millipore) (Schmued et al., 2005 (link)) and 2 μM DAPI in a 0.1% (v/v) acetic acid, and rinsed by three washes with distilled water. Sections were dried at 50 °C, dehydrated in xylene, and mounted with DPX Mountant (Sigma-Aldrich). Images of the hippocampal region were captured with a Leica TCS SP8 laser-scanning microscope equipped with a 10 × objective. To quantify Fluoro-Jade C-positive cells, four mice per condition were used. For each hippocampus, six sections were analyzed. Neuronal degeneration was expressed as the intensity of CA3 fluorescence divided by that of CA1 fluorescence in the pyramidal cell layer.
5
Histochemical Staining Protocols for Tissue Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
H&E staining was carried out according to the following method. After deparaffinization, sections were stained with Mayer's hematoxylin solution (Wako, Tokyo, Japan) for 10 min, and then stained with eosin alcohol solution (Wako) for 4 min. Finally, the sections were dehydrated and coverslipped using NEW M.X (Matsunami-glass). FluoroJade C (#3,319,822; Merck Millipore, Burlington, MA, USA) staining was carried out according to the manufacturer’s instructions. Briefly, frozen sections were treated with basic ethanol (80% ethanol/1% NaOH) for 5 min, and washed in 70% EtOH and ultrapure water for 2 min, respectively. The sections were then incubated in 0.06% potassium permanganate. After washing in ultrapure water for 2 min, the sections were treated with the staining solution (Fluoro-Jade C and 2 µg/mL Hoechst diluted in 0.1% acetic acid). After washing in ultrapure water, the sections were heated at 60 °C, air dried, and incubated in xylene for 5 min. Finally, the sections were coverslipped using NEW M.X (Matsunami-glass). Floro-Styryl-Benzene (FSB) (Dojindo, Tokyo, Japan) staining was carried out according to the manufacturer's instructions. The sections were then incubated with FSB solution, and then soaked in lithium carbonate solution. After washing with 50% EtOH, the sections were coverslipped.
6
Fluoro-Jade C Staining for Neuronal Degeneration
7
Fluoro-Jade C Protocol for Neurodegeneration
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The tissue cryostat sections were mounted on Superfrost Plus microscope slides (FisherScientific, 22–230-892) and dried at 50 °C for 30 min. The slides were immersed in the following solutions at room temperature in order: 0.2% NaOH in 80% ethanol for 5 min, 70% ethanol for 2 min, distilled water for 2 min, 0.06% potassium permanganate for 10 min, distilled water for 2 min, 0.0001% Fluoro-Jade C (Sigma-Aldrich, AG325) in 0.1% acetic acid for 10 min and 3 times distilled water for 1 min. The slides were dried at 50 °C for 10 min, immersed in xylene for 1 min and coverslipped with DPX Mounting medium (Millipore Sigma, 06522).
For double labeling, regular Immunofluorescence protocol as described above was performed first, followed by the Fluoro-Jade staining protocol.
For double labeling, regular Immunofluorescence protocol as described above was performed first, followed by the Fluoro-Jade staining protocol.
8
Fluoro-Jade C Staining for Neurodegeneration
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tissue was fixed in formalin, cryoprotected using a solution of 200 g/L glucose in PBS, and frozen at −20°C for cross‐sectioning at 20 µm. Prior to staining, sections were mounted onto slides and air‐dried for 20 min. Slides were first immersed in a basic alcohol solution consisting of 10 g/L sodium hydroxide in 800 ml/L ethanol for 5 min, followed by two rinses of 2 min in 700 ml/L ethanol and 2 min in distilled water. Slides were then incubated in 60 mg/L potassium permanganate solution for 15 min. Following a 2‐min water rinse, slides were transferred for 15 min to a 1 mg/L solution of Fluoro‐Jade C (Sigma‐Aldrich, Germany) in 1 ml/L acetic acid vehicle. The slides were then rinsed through three 1‐min changes of distilled water. Excess water was drained onto a paper towel, and the slides were air‐dried for 10 min. Slides were then mounted with VECTASHIELD Mounting Medium with DAPI (Vector Laboratories, USA).
9
Fluoro-Jade C Protocol for Neurodegeneration
10
Fluoro-Jade C Staining for Neurodegeneration
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Experimental procedures for preparing frozen sections were the same as for immunofluorescence staining. FJC staining was performed using a standard protocol. Briefly, brain tissue sections were immersed in 1% sodium hydroxide in 80% ethanol for 5 minutes, then rinsed for 2 minutes in 70% ethanol, for 2 minutes in distilled water, followed by incubating in 0.06% potassium permanganate solution for 10 minutes. After a 1–2 minute water rinse, the slides were transferred to a 0.0001% solution of Fluoro-Jade C (Sigma Aldrich, USA) dissolved in 0.1% acetic acid vehicle for 10 minutes. The slides were then rinsed through three changes of distilled water for 1 minute per change, then air dried on a slide warmer at 50°C for at least 5 minutes. Last, sections were immersed in xylene for 1 minute and mounted with DPX.
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!