HeLa cells treated with rotenone or MnCl2 were assessed for cell death induction as previously described [48 (link)]. Briefly, treated cells were collected at 48 h by trypsinization, PBS-washed and exposed to 1 µg/ml propidium iodide (PI; Sigma-Aldrich, P4170-25MG) for 5 min. PI positivity was captured using an Attune flow cytometer (Life Technologies). Neuronal cell apoptosis was assessed by cleaved caspase 3 staining in accordance with the manufacturer’s guidelines (Cell Signaling, 9661). Cortical neurons were plated with or without cover slips. Cells plated without coverslips were ultimately assessed by flow cytometry using an Attune flow cytometer. Samples seeded on coverslips were imaged using a LSM780 confocal microscope (Leica). DAPI staining was used to visualize the nucleus.
P4170 25mg
Manufactured by Merck Group
P4170-25MG is a chemical compound produced by Merck Group. It is available in a 25-milligram quantity. No further details about its core function or intended use can be provided in an unbiased and factual manner.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 780, by Zeiss (2 mentions)
Attune flow cytometer, by Thermo Fisher Scientific (1 mentions)
Lsm780 confocal microscope, by Leica (1 mentions)
B2261 25mg, by Merck Group (1 mentions)
Eclipse te300, by Nikon (1 mentions)
Hoechst 33342, by Merck Group (1 mentions)
Mab386, by Merck Group (1 mentions)
Flouromount g, by Southern Biotech (1 mentions)
Alexa fluor 633 goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Alexa fluor 488 goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Tcs sp2, by Leica (1 mentions)
5 protocols using p4170 25mg
1
Assessing Cell Death and Apoptosis
2
Whole-Genome Sequencing of Evolved Isolates
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Whole-genome sequencing was performed for 30 evolved isolates from different subpopulations at the early time point, and the three subpopulations of #8 at the late time point (SI Appendix ). For defined mutants (SI Appendix ), we assessed the change in growth upon pretreatment by OD600 every 15 min. We additionally used flow cytometry (Guava EasyCyte HT Blue-Green; Merck KGaA) with hourly samples, Live/Dead staining, and three technical replicates. For staining, cells were incubated for 10 min with propidium iodide (P4170-25MG; Sigma-Aldrich) and thiazole orange (390062-250MG; Sigma-Aldrich). We assessed antibiotic tolerance for isolate 12-1a-E2-4 (isolated after transfer 12 from sequence #12) via minimal duration of killing (25 (link)).
3
Seedling Microscopic Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Seedlings were mounted with distilled water and observed using either an epifluorescence microscope (Olympus BX-21) or a confocal microscope (Zeiss LSM 780). Cell walls and nuclei were visualized by staining with propidium iodide (PI, SIGMA, P4170-25MG, 30 μM in distilled water) for 10 minute and samples were mounted with ddw. To quantify DCFDA intensities, all fluorescence images were taken at the same exposure time. DCFDA signal intensities were quantified using ImageJ with mean grey value options by measuring data within whole cotyledon region. Numbers of cotyledon to measure the DCFDA signal intensity were indicated in each figure legends. PI signal was gained between 600–630 nm, and chlorophyll autofluorescence signal was gained between 600–780 nm.
4
Blastocyst Evaluation and BOEC Viability
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Blastocyst rates were evaluated at day 7 and 8 of embryo culture and defined as the total number of blastocysts on day 7 or 8 divided by the total number of oocytes selected for maturation that were fertilized. The viability of BOEC after the co-culture was assessed by double staining with Propidium Iodide (PI, P4170-25MG, Sigma) and Hoechst 33342 (B2261-25MG, Sigma) and observed under fluorescent microscope (Nikon Eclipse TE 300) [27 (link)].
5
Confocal Imaging of Nerve Fiber and Macrophage
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Confocal laser microscopy was used to analyze the distribution of CD163 labelled macrophages (method see below) and neurofilament 200 (NF) and myelin basic protein (MBP), markers of axon and myelin sheath, respectively, in peripheral nerve specimens. CD 163 labeling was done on nerve sections from implanted, proximal, distal nerves as well as on nerve samples before implantation. Samples were incubated with mouse anti-CD163 antibody (1:200, Novocastra NCL-L-CD163, Clone 10D6) followed by incubation with Alexa Fluor 488 goat anti-mouse IgG (1:200, Invitrogen A11001), while nuclei were stained with Propidium iodide (1:3000, Sigma-Aldrich P4170-25 MG). For evaluation of the proportion of myelinated nerve fibers, specimens were first incubated with monoclonal mouse anti-NF antibody (1:75, Dako M0762, Clone 2F11), followed with Alexa Fluor 488 goat anti-mouse IgG (1:200, Invitrogen A-11001) and later incubated with monoclonal rabbit anti-MBP antibody (1:50, Millipore MAB386), followed with Alexa Fluor 633 goat anti-rabbit IgG (1:200, Invitrogen A21070). Finally, sections were mounted with Flouromount-G (SouthernBiotech, 0100-01) and images were acquired with Leica Confocal Software on Confocal Scanner microscope Leica TCS SP2 (Leica Microsystems, Wetzlar, Germany).
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!