Immunohistochemistry was carried out as previously reported with minor modifications (Aricioglu et al., 2019 (link)). Brain tissue was fixed with 4% paraformaldehyde for 24 h. Dehydrated, embedded in paraffin, and cut into slices of 5 µm thickness. After a series of operations, including deparaffinization, antigen repair and blocking, sections were incubated overnight with primary antibodies: anti-BDNF (Abcam, ab108319, 1:200), anti-P2X7 (Proteintech, 28207-1-AP, 1:50), anti-NLRP3 (Abcam, ab214185, 1:200) and anti-Cleaved caspase-1 (Thermo Fisher, PA5-99390, 1:200) at 4°C, followed by secondary antibodies incubation. Then, the slices were colored with diaminobenzidine (DAB). Finally, after dehydration and drying, sections were observed using a light microscope. The density values are analyzed by Image pro-plus software.
Pa5 99390
Manufactured by Thermo Fisher Scientific
The PA5-99390 is a lab equipment product manufactured by Thermo Fisher Scientific. It is designed to perform a core function within the laboratory setting. However, a detailed and unbiased description of its features and capabilities is not available at this time.
Automatically generated - may contain errors
Lab products found in correlation
Ab214185, by Thermo Fisher Scientific (1 mentions)
Anti cleaved caspase 1, by Thermo Fisher Scientific (1 mentions)
Ab108319, by Proteintech (1 mentions)
Ab219800, by Abcam (1 mentions)
Ab214185, by Abcam (1 mentions)
Ab175449, by Abcam (1 mentions)
Ab234437, by Abcam (1 mentions)
Ab223694, by Abcam (1 mentions)
Ab191860, by Abcam (1 mentions)
Ab133514, by Abcam (1 mentions)
Ab6728, by Abcam (1 mentions)
3 protocols using pa5 99390
1
Immunohistochemical Analysis of Neuroinflammation Markers
2
Western Blot Analysis of NLRP3 Inflammasome
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lysates from tissues and cells were prepared using RIPA lysis buffer and were quantified using a BCA kit (cat. #BCA1; Sigma–Aldrich). Afterwards, lysates were resolved in SDS-PAGE to separate proteins and were then transferred to polyvinylidene fluoride membranes. The membranes were immersed in 5% skimmed milk to block for non-specific binding and labelled with anti-WHSC1 (1:2000; cat. #ab223694; Abcam), NLRP3 (1:500; cat. #ab214185; Abcam), ASC (1:2000; cat. #ab175449; Abcam), caspase-1 p20 (1:1000; cat. #PA5-99390; Thermo Fisher Scientific), IL-18 (1:1000; cat. #ab191860; Abcam), IL-1β (1:1000; cat. #ab234437; Abcam), GSDMD FL (full length GSDMD; 1:1000; cat. #ab219800; Abcam), GSDMD CL (GSDMD-N terminal segment; 1:1000; cat. #10137S; Cell Signaling Technology, Danvers, MA), NEK7 (1:3000; cat. #ab133514; Abcam) and β-actin (1:5000; cat. #ab8227; Abcam) Abs at 4°C for 12 h. Afterwards, the membranes were incubated with IgG H&L (HRP; 1:5000; cat. #ab6728; Abcam) for 2 h at room temperature. β-Actin was designated as the internal control protein. Densitometry of bands was measured utilising the ECL Western Blotting Substrate (cat. #32132X3; Thermo Fisher Scientific).
3
Immunohistochemical Detection of Cleaved Caspase-1
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The expression patterns of cleaved caspase-1 in cartilage tissues were detected by referring to methods in previous literature [19 (link)]. Briefly, the tissues were fixed with 4% paraformaldehyde, paraffin-embedded, sectioned continuously (at 5 μm), baked at 58°C for 18 h, and dewaxed with xylene. Next, the sections were treated with 0.01 mol/L citric acid buffer at 95°C (5 min × 2 times) for antigen retrieval, incubated with 3% H2O2 for 15 min at room temperature, and then incubated with the primary antibody (cleaved caspase-1, dilution ratio of 1:1000, PA5-99,390, Thermo Fisher Scientific) at 4°C overnight and the secondary antibody at room temperature for 30 min. Afterward, the sections were developed with 2,4-diaminobutyric acid for 5 min, rinsed under running water to terminate the reaction, counterstained with hematoxylin, and sealed with resin. Subsequently, 9 visual fields were randomly selected from each sample to calculate the percentage of 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!