Brain tissues were sectioned on a standard rotary microtome (Leica, Berlin, Germany). Tissue sections were collected and counterstained with hematoxylin-eosin (HE). Then, the sections were subjected to staining with the primary antibodies. Primary antibodies were tyrosine hydroxylase (TH, 1:1000) (Santa Cruz Biotech., Dallas, TX, USA), P-CREB (1:500, Upstate Comp., Lake Placid, NY, USA), caspase 3 (1:500), or MDA (1:100). After raining with PBS, sections were stained with 3,3′-diaminobenzidine (DAB) as described [24 (link)]. Subsequently, the selected sections were further performed by using Tuj1 (1:500), GFAP (1:500), and IBA1 antibody (1:200, Santa Cruz Biotech., Dallas, TX, USA). The stained sections were counterstained with DAPI, mounted and examined microscopically.
Standard rotary microtome
Manufactured by Leica
Sourced in Germany
The Standard rotary microtome is a precision instrument used to cut thin sections of samples for microscopic examination. It features a manual rotary mechanism that precisely controls the thickness of the sliced sections, allowing for consistent and accurate sample preparation.
Automatically generated - may contain errors
Lab products found in correlation
Glial fibrillary acidic protein (gfap), by Santa Cruz Biotechnology (1 mentions)
Tuj 1, by Santa Cruz Biotechnology (1 mentions)
Tyrosine hydroxylase th, by Santa Cruz Biotechnology (1 mentions)
Iba1 antibody, by Santa Cruz Biotechnology (1 mentions)
Light microscopy, by Nikon (1 mentions)
Ab269452, by Abcam (1 mentions)
Dp12 digital camera, by Olympus (1 mentions)
4 protocols using standard rotary microtome
1
Brain Tissue Immunohistochemistry Protocol
2
Histological Analysis of Tissue Samples
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For histological analysis, tissues have been explanted and immediately fixed overnight in 4% formaldehyde at 4°C. They have been dehydrated and paraffin-embedded. Five-micrometer-thick sections have been generated using standard rotary microtome (Leica) and stained using hematoxylin and eosin or Picrosirius red. For immunohistochemistry, we used proteinase K–mediated antigen unmasking or heat-induced antigen unmasking using tris-EDTA (pH 8). Mouse-on-mouse blocking reagent (Abcam, ab269452) or 5% normal goat serum has been used before incubation with primary antibodies. The list of primary and secondary antibodies used has been provided (table S2). Images have been collected using light microscopy (Nikon). Immunofluorescence images were analyzed with a custom Fiji Macro through a nuclei segmentation and a dot analysis. Mean gray value (the sum of the gray values of all the pixels divided by the number of the pixels) is used to graph the results obtained.
3
Tissue Sectioning and HE Staining
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tissue blocks were sectioned at 4–5 μm on a standard rotary microtome (Leica, Germany). These sections were collected and then stained by HE. The stained sections were mounted and examined microscopically (Nikon, NI-E).
4
Histological Analysis of Larvae
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The same larvae used in the tracking studies were subsequently used for the histological analysis. In order to avoid excessive body bending during processing, larvae were fixed and dehydrated inside 1 cm diameter glass tubes. Thus, larvae were fixed in 1% phosphate-buffered paraformaldehyde (pH 7.3) overnight at 4ºC. After rinsing in PBS 1x, samples were dehydrated with increasing concentrations of ethanol (70, 90, and 96%), cleared in butanol, and embedded in low melting paraffin (56-58ºC, Lab-o-wax, Histo-line Laboratories, Pantigliate, Italy).
Paraffin blocks were sectioned at 10 μm on a standard rotary microtome (Leica, Wetzlar, Germany). These sections were collected on gelatinized slides and then stained by hematoxylin eosin (HE) following standard protocols. The stained sections were mounted in Eukitt ® and examined microscopically in an optical microscope (BX61, Olympus ® , Tokyo, Japan) equipped with a DP12 digital camera (Olympus ® , Tokyo, Japan). Low magnification images were obtained using an MPlan Apo 1.24/0.04 objective (Olympus ® , Tokyo, Japan).
Paraffin blocks were sectioned at 10 μm on a standard rotary microtome (Leica, Wetzlar, Germany). These sections were collected on gelatinized slides and then stained by hematoxylin eosin (HE) following standard protocols. The stained sections were mounted in Eukitt ® and examined microscopically in an optical microscope (BX61, Olympus ® , Tokyo, Japan) equipped with a DP12 digital camera (Olympus ® , Tokyo, Japan). Low magnification images were obtained using an MPlan Apo 1.24/0.04 objective (Olympus ® , Tokyo, Japan).
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!