The tissue samples surrounding first molar were homogenized and then lysed using Protein Lysis Buffer (Promega Corporation). Protein concentration was measured using bicinchoninic acid protein assay method (Nanjing Jiancheng Bioengineering Institute). The same amount of protein (30 µg) was separated using 10% SDS-PAGE. The proteins were transferred to a nitrocellulose membrane (EMD Millipore). Following blocking with 5% non-fat milk prepared with TBS-Tween 20 (0.1%) at room temperature for 2 h, the membranes were incubated with the primary antibodies aforementioned overnight at 4˚C. After washing, the membranes were incubated with the horseradish peroxidase-conjugated secondary antibodies (Goat anti-rabbit IgG; cat. no. ab97051; 1:2,000; Goat anti-mouse IgG, cat. no. ab205719, 1:2,000; Abcam) aforementioned for 2 h at room temperature. SuperSignal™ West Femto Maximum Sensitivity Substrate (Thermo Fisher Scientific, Inc.) was used as visualization reagent. ImageJ software 1.53 version (National Institutes of Health) was used to analyze the protein band intensity.
Protein lysis buffer
Manufactured by Promega
Sourced in United States
Protein Lysis Buffer is a solution designed to facilitate the extraction and solubilization of proteins from biological samples. It is a key component in the sample preparation process for various protein analysis techniques.
Automatically generated - may contain errors
Lab products found in correlation
Supersignal west femto maximum sensitivity substrate, by Thermo Fisher Scientific (1 mentions)
Ab81484, by Abcam (1 mentions)
Oleate, by Merck Group (1 mentions)
Dual luciferase reporter dlr assay system, by Promega (1 mentions)
A939572, by Merck Group (1 mentions)
Glomax 20 20 luminometer, by Promega (1 mentions)
Pro200 series homogenizer, by PRO Scientific (1 mentions)
Protease inhibitor cocktail, by Roche (1 mentions)
3 protocols using protein lysis buffer
1
Western Blot Analysis of Protein Targets
2
Inhibition of Wnt Signaling Pathway
3
Cytokine and Chemokine Profiling in Skin Infections
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Infected skin biopsies from day 3 were analyzed for protein levels of cytokines, chemokines and growth factors to provide insights into the host response to CA-MRSA or LA- S. aureus i.d. inoculation. Mice (n = 5/group) were euthanized on day 3 post-infection and 10-mm skin punch biopsies of lesions were weighed and snap-frozen in liquid nitrogen. On ice, each specimen was homogenized with a hand-held homogenizer (Pro200 Series homogenizer; Pro Scientific) in Protein Lysis Buffer (Promega) containing protease inhibitor cocktail (Roche). All samples were stored at 80 °C. Samples were then centrifuged at 4 °C and supernatants were assayed for protein levels of cytokines, chemokines and growth factors using a 9-plex and 11-plex mouse protein array, according to the manufacturer’s recommendations (Bio-Plex Pro™, Biorad; Hercules, CA). For the 9-plex and 11-plex arrays, samples were normalized to 0.75 mg/mL and 2 mg/mL total protein, respectively. Samples were also assayed for myeloperoxidase (MPO) levels using a commercially available ELISA kit (R&D Systems, Minneapolis, MN).
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!