The hippocampus from the other half of the brain (n = 6 for each group) was isolated and homogenized for biochemical assays. Equal amounts of protein (35 μg/lane) were electrophoretically separated and blotted onto nitrocellulose membranes. Protein levels were determined via incubation against antibody of NLRP3 (1:600; Servicebio Technology Co., Ltd.), ASC (1:200; Santa Cruz, CA, United States), caspase-1 (1:500; Servicebio Technology Co., Ltd.), gasdermin-D (GSDMD, 1:500; Abcam, United Kingdom), IL-1β (1:200; Santa Cruz, United States), IL-18 (1:200; Santa Cruz, CA, United States), or β-actin (1:1,000, Cell Signaling Technology, United States). Bands were visualized by enhanced chemiluminescence and quantified with the Image Quant Software (Syngene).
Gasdermin d gsdmd
Manufactured by Abcam
Sourced in United Kingdom
Gasdermin D (GSDMD) is a protein that plays a key role in the process of pyroptosis, a form of programmed cell death. It is a member of the gasdermin family of proteins, which are involved in the regulation of inflammatory responses. GSDMD is known to be involved in the formation of pores in the cell membrane, leading to cell lysis and the release of inflammatory factors.
Automatically generated - may contain errors
Lab products found in correlation
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Image quant software, by Syngene (1 mentions)
Il 1β, by Santa Cruz Biotechnology (1 mentions)
Caspase 1, by Wuhan Servicebio Technology (1 mentions)
Nlrp3, by Wuhan Servicebio Technology (1 mentions)
Il 18, by Santa Cruz Biotechnology (1 mentions)
β actin, by Cell Signaling Technology (1 mentions)
Il 1b, by Abcam (1 mentions)
Il 18, by Solarbio (1 mentions)
Phospho stat3 tyr705, by Cell Signaling Technology (1 mentions)
Caspase 1, by Huabio (1 mentions)
Total jak2, by Cell Signaling Technology (1 mentions)
Total stat3, by Cell Signaling Technology (1 mentions)
Cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Bcl2 associated x (bax), by Proteintech (1 mentions)
Bcl 2, by Proteintech (1 mentions)
Pro caspase 1, by Abcam (1 mentions)
Cleaved caspase 1, by Abcam (1 mentions)
Gsdmd n, by Abcam (1 mentions)
C ebp homologous protein chop, by Abcam (1 mentions)
4 protocols using gasdermin d gsdmd
1
Hippocampal Inflammatory Protein Analysis
2
Hippocampal Protein Analysis after HSR
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
At 12 h after HSR, total protein was extracted from the hippocampal tissues (n=6 per group) as described in the IL-1β and IL-18 assay. Samples containing 30 µg protein quantified by bicinchoninic acid (BCA) assay were separated by 10% SDS-PAGE and transferred onto a PVDF membrane. The membrane was blocked with 5% skimmed milk at 37°C for 2 h, and primary antibodies rabbit anti-rat polyclonal cleaved caspase-1 (1:1,000; cat. no. ab1872; Abcam), monoclonal NLRP3 (1:1,000; cat. no. ab210491; Abcam), polyclonal Gasdermin D (GSDMD; 1:1,000; cat. no. ab219800; Abcam), polyclonal IL-1b (1:1,000; cat. no. K107559P; Beijing Solarbio Science & Technology Co., Ltd.) and polyclonal IL-18 (1:1,000; cat. no. K002143P; Beijing Solarbio Science & Technology Co., Ltd.) were applied at 4°C overnight, followed by incubation with a horseradish peroxidase-conjugated goat anti-rabbit IgG secondary antibody (1:2,000; cat. no. sc-2004; Santa Cruz Biotechnology, Inc.) at 25°C for 1 h. Protein bands in each treatment group were detected using an enhanced chemiluminescence western blot detection system ImageJ 1.48u software (National Institutes of Health). β-actin (1:2,000; cat. no. sc-47778; Santa Cruz Biotechnology, Inc.) was used as an internal reference (20 (link)).
3
Protein Expression Analysis in HK-2 Cells and Mice Kidneys
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total proteins of cultured HK-2 cells and kidneys of the mice were extracted with cell lysis buffer supplemented with protease and phosphatase inhibitor cocktails. The concentration of proteins was detected with a BCA Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA). Equal amounts of proteins were separated by 10% sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidene fluoride membranes. After being blocked with 5% skim milk for 1 h, the membranes were incubated overnight at 4°C with antibodies directed against total STAT3 and phospho-STAT3-Tyr705 (1:2,000, Cell Signaling Technology), total JAK2 (1:1,000, Cell Signaling Technology), phospho-JAK2-Tyr1007/1008 (1:2,000, Abcam), cleaved caspase-3 (1:1,000, Cell Signaling Technology), Bax (1:2,000, ProteinTech), Bcl-2(1:1,000, ProteinTech), gasdermin D (GSDMD, 1:1,000, Abcam), and caspase-1 (1:2,000, Huabio). The next day, the membranes were washed and incubated with horseradish peroxidase-conjugated secondary antibodies for 1 h at room temperature. Protein bands were visualized using enhanced chemiluminescence substrate and quantified by ImageJ software ver. 1.8.
4
LPS-Induced Pyroptosis Mechanism
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
We detected the role of UA in cell pyrophosis. We observed that LPS stimulated the production of NLRP3, The supernatants were collected through centrifugation and subjected to protein concentration determination with bicinchoninic acid (BCA) protein assay kit (Beyotime Biotechnology, Shanghai, China). Proteins were resolved with 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred onto polyvinylidene difluoride (PVDF) membranes. Membranes were then blocked with 5% bovine serum albumin (BSA) followed by incubation with primary antibodies targeting LC3-II/I (1:1000; Thermo Scientific, Waltham, MA, USA), Beclin (1:1000; Abcam, Cambridge, UK), p62 (1:1000; Abcam), gasdermin D (GSDMD, 1:1000; Abcam), GSDMD-N (1:1000; Abcam), NLR family pyrin domain containing 3 (NLRP3, 1:1000; Abcam), pro-caspase-1 (1:1000; Abcam), cleavedcaspase-1 (1:1000; Abcam), pro-IL-1β (1:1000; Abcam), cleaved IL-1β (1:1000; Abcam), activating transcription factor 6 (ATF6, 1:1000; Abcam), C/EBP-homologous protein (CHOP, 1:1000; Abcam), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH, 1:10000; Abcam). Membranes were incubated with HRP-conjugated secondary antibodies at a ratio of 1:1000 for 2 h after rinsing in tris buffered saline with tween (TBST) for 15 min. The signals were detected with enhanced chemiluminescence (ECL) detection kit.
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!