Fresh lung tissue was fixed with 4% paraformaldehyde (PFA), dehydrated in a sucrose solution gradient, wrapped with Tissue-Tek OCT, and sectioned into 5-μm slices. In addition, PMNs or NRR8383 cells were seeded over a 14-mm coverslip (Solarbio, Beijing) in 24-well plates overnight. Immediately afterwards, cell and tissue slices were fixed in 4% PFA, permeabilized, blocked with 5% goat serum to minimize nonspecific staining, and incubated with the following primary antibodies overnight at 4 ℃. Rabbit polyclonal anti-citrullinated histone H3 (cit-H3, 1:250, Abcam, UK), rat polyclonal anti-LY6G (1:150, Abcam, UK), mouse monoclonal anti-ASC (1:100, Santa Cruz Biotechnology, USA), mouse polyclonal anti-CD68 (1:200, Immunoway Biotechnology, UK), rabbit polyclonal anti-NLRP3 (1:200, Boster, China) and mouse monoclonal anti-ubiquitin (1:100, Santa Cruz Biotechnology, USA) antibodies. Following incubation with secondary Alexa Fluor 488 (1:200, Elabscience, China) and Alexa Fluor 594 (1:200, Elabscience, China) for 1 h at room temperature. The nucleus was stained with 4′,6-diamidino-2-phenylindole (DAPI, Boster, China) for 5 min, protected from light. In addition, NETs-associated DNA was detected with SYTOX green in accordance with the manufacturer’s specifications. Samples were visualized by confocal laser scanning microscopy (Olympus) or fluorescence microscopy (Zeiss).
Alexa fluor 488
Manufactured by Elabscience
Sourced in China
Alexa Fluor 488 is a fluorescent dye manufactured by Elabscience. It is a green-fluorescent dye with excitation/emission maxima of approximately 488/519 nm. Alexa Fluor 488 can be used for various fluorescence-based applications.
Automatically generated - may contain errors
Lab products found in correlation
Fluorescence microscopy, by Zeiss (1 mentions)
Anti ly6g, by Abcam (1 mentions)
Anti cd68, by Immunoway (1 mentions)
Anti nlrp3, by Boster Bio (1 mentions)
Mouse monoclonal anti ubiquitin, by Santa Cruz Biotechnology (1 mentions)
4 6 diamidino 2 phenylindole dapi, by Boster Bio (1 mentions)
Confocal laser scanning microscopy, by Olympus (1 mentions)
Related kits, by Pars Azmoon (1 mentions)
Cdna kit, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
Elisa kit, by ZellBio (1 mentions)
Cyanine3 (cy3), by Elabscience (1 mentions)
Mouse anti 2 3 cyclic nucleotide 3 phosphodiesterase cnpase, by Abcam (1 mentions)
Dm6b fluorescence microscope, by Leica camera (1 mentions)
3 protocols using alexa fluor 488
1
Immunofluorescence Analysis of Lung Tissue and Cells
2
Characterization of Alumina Nanoparticles and Biomarkers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The α-Al2O3 NPs (80 nm, 100% alpha, white, hydrophilic, purity 99%, SSA: > 15 m2/g, density 3.97 g/cm3 with rhombohedral crystallographic structure) and γ-Al2O3 NPs (20 nm, white, 99% purity, SSA: > 138 m2/g, density 3890 kg/m3, almost spherical morphology) were purchased from US Research Nanomaterials Inc., Houston, TX USA (CAS No 1344–28-1, USA). Additionally, all the ultra-pure chemicals used in this research were purchased from Merck Company. MDA (CAS No. ZB-MDA-A96A), GPX (CAS No. ZB-GPX-A96), T-SOD (CAS No. 706002), TAC (CAS No. ZB-TAC-A96) CAT (CAS No. 707002), iNOS (CAS No. ZB-10740C-R9648) were assessed using ELISA kits (Zellbio Germany) and cDNA kit (Thermo scientific, US, K1622). For immunohistochemical assays, the following reagents were used: TBS 1X solution (Sigma-T5912), PBS (Sigma-P4417), DAPI (Sigma- D9542) Normal Goat Serum (10%) (G9023-Sigma), Triton 3% (Sigma-T8787), Alexa Fluor 488 (Elabscience, China), CYP450: GTX15616, Secondary antibodies (mouse): orb688924. SYBR Green master mix (Addbio Co., Korea) was utilized, following specific standards for each marker. AST and ALT were assessed using related kits (Pars Azmun, Tehran, Iran).
3
Immunofluorescence Staining of Spinal Cord Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The fixed NSCs and a 4-μm-thick longitudinal slice of spinal cord (centered on the epicenter of the injured lesion) were prepared for immunofluorescence staining as described in our previous study [42 (link)]. The primary antibodies were used as follows: mouse anti-2′3′ cyclic nucleotide 3' phosphodiesterase (Cnpase,1:200; Abcam, UK) and rabbit anti-myelin basic protein(MBP, 1:300; Abcam, UK) for oligodendrocytes, rabbit anti-glial fibrillary acidic protein (GFAP) for astroglia (1:1000; Abcam, UK), mouse anti-nestin for NSCs (1:1000; Abcam, UK), rabbit anti-neuron-specific class III beta-tubulin (Tuj1) for neuron (1:1000; Abcam, UK),rabbit anti- SRY-Box transcription factor 10 (Sox 10) for NSCs (1:100; Abcam, UK), rabbit anti- p75 neurotrophin receptor (p 75) for NSCs (1:100, Abcam, UK), and rabbit anti-BMP2 (1:500; Affinity, USA). The secondary antibodies used were Cy3 (red, 1:50; Elabscience, China) and Alexa Fluor 488 (green, 1:50; Elabscience, 288 China). The images were observed and photographed by using a DM-6B fluorescence microscope (Leica, Germany). The percentage of positive areas was calculated using ImageJ. For cell counting, random fields containing a total of 500 cells were randomly selected. The number of positive cells was blindly quantitated in two different individuals.
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!