The transwell assay was performed as described previously34 (link). Using transwell inserts (No 354480; Corning; NY; 14831), a transwell assay was performed in 5 groups: (1) the control group: primary astrocytes in the inserts, with RPMI 1640 in the well; (2) the macrophage group: primary astrocytes in the inserts, with RPMI 1640 with primary macrophages in the well; (3) the ADP group: primary astrocytes in the inserts, with RPMI 1640 with ADP (100 μM; Oriental Yeast, Tokyo, Japan) in the well; (4) the apyrase group: primary astrocytes in the inserts, with RPMI 1640 with primary macrophages and apyrase (10 U/ml; Sigma, Saint Louis, MO) in the well; and (5) the MRS-2179 group: primary astrocytes treated with MRS-2179 (10 μM; Abcam, Cambridge, UK) in the inserts, with RPMI 1640 with primary macrophages in the well. Each insert was seeded with 5.0 × 104 cells of astrocytes and each well was seeded with 5.0 × 104 cells of macrophages. After incubation for 24 h and staining with Diff-Quick (Sysmex, Kobe, Japan), the number of migrating cells was counted in 9 sections/3 wells per group. The percentage increase in migrating cells compared to the control group was evaluated.
Mrs 2179
Manufactured by Abcam
Sourced in United Kingdom
MRS-2179 is a selective P2Y1 receptor antagonist. It is commonly used in research to investigate the role of the P2Y1 receptor in various biological processes.
Automatically generated - may contain errors
Lab products found in correlation
Apyrase, by Merck Group (2 mentions)
Adenosine diphosphate (adp), by Oriental Yeast (2 mentions)
Rpmi 1640, by Oriental Yeast (2 mentions)
Rpmi 1640, by Merck Group (2 mentions)
Transwell insert, by Corning (2 mentions)
Diff quick, by Sysmex (2 mentions)
Kds 310, by Muromachi Kikai (1 mentions)
Hexamethonium, by Merck Group (1 mentions)
Nicardipine, by Merck Group (1 mentions)
Hyoscine, by Merck Group (1 mentions)
Calcein am, by Santa Cruz Biotechnology (1 mentions)
Tnf α, by Thermo Fisher Scientific (1 mentions)
Ar c66096, by Bio-Techne (1 mentions)
Mrs2578, by Bio-Techne (1 mentions)
Bms ccr2 22, by Bio-Techne (1 mentions)
Arl67156, by Bio-Techne (1 mentions)
Fluo 4 am, by Thermo Fisher Scientific (1 mentions)
Bapta am, by Bio-Techne (1 mentions)
Mrs2211, by Abcam (1 mentions)
5 protocols using mrs 2179
1
Transwell Assay for Astrocyte-Macrophage Interactions
2
Transwell Assay for Astrocyte Migration
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The transwell assay was performed as described previously25 (link). Using transwell inserts (No 354480; Corning; NY; 14831), a transwell assay was performed in 5 groups: (1) the control group: primary astrocytes in the inserts, with RPMI1640 in the well; (2) the macrophage group: primary astrocytes in the inserts, with RPMI 1640 with primary macrophages in the well; (3) the ADP group: primary astrocytes in the inserts, with RPMI 1640 with ADP (100μM; Oriental Yeast, Tokyo, Japan) in the well; (4) the Apyrase group: primary astrocytes in the inserts, with RPMI 1640 with primary macrophages and Apyrase (10 U/ml; Sigma, Saint Louis, MO) in the well; and (5) the MRS-2179 group: primary astrocytes treated with MRS-2179 (10μM; Abcam, Cambridge, UK) in the inserts, with RPMI 1640 with primary macrophages in the well. After incubation for 24 hours and staining with Diff-Quick (Sysmex, Kobe, Japan), the number of migrating cells was counted in 9 sections/3 wells per group. The percentage increase in migrating cells compared to the control group was evaluated.
3
Spinal Cord Injury Treatment
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
4
Pharmacological Modulation of Enteric Neuron-Driven Motility
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The results support the idea that cell therapy might be used to treat motility disorders caused by missing or diseased enteric neurons. 100 ms was chosen because it consistently evoked responses in the muscle large enough to allow pharmacologic studies (Supplementary Figure 1).
We recorded 5À8 inhibitory or excitatory responses in the circular muscle cells evoked by each of electrical stimulation or light stimulation before addition of antagonists to the bath solution. A similar number of responses to each type of stimulation was recorded in the presence of antagonists before the reversibility of antagonists was tested after a wash period of at least 15 minutes. If the impalement were lost during recordings, a new impalement was made at the same location, taking advantage of the syncytial nature of the smooth muscle. Drugs used in the experiments were hyoscine, hexamethonium, nicardipine, pyridoxal phosphate-6-axophenyl-2 0 -4 0disulfonic acid (PPADS), nitro-L-arginine (NOLA), trinitrophenyladenosine 5 0 -triphosphate (TNP-ATP) (all from Sigma Aldrich, Castle Hill, NSW, Australia), and MRS 2179 (Abcam Biochemicals, Melbourne, Australia). Stock solutions of antagonists were initially made up in distilled water and diluted to working concentrations on the day of the experiment.
We recorded 5À8 inhibitory or excitatory responses in the circular muscle cells evoked by each of electrical stimulation or light stimulation before addition of antagonists to the bath solution. A similar number of responses to each type of stimulation was recorded in the presence of antagonists before the reversibility of antagonists was tested after a wash period of at least 15 minutes. If the impalement were lost during recordings, a new impalement was made at the same location, taking advantage of the syncytial nature of the smooth muscle. Drugs used in the experiments were hyoscine, hexamethonium, nicardipine, pyridoxal phosphate-6-axophenyl-2 0 -4 0disulfonic acid (PPADS), nitro-L-arginine (NOLA), trinitrophenyladenosine 5 0 -triphosphate (TNP-ATP) (all from Sigma Aldrich, Castle Hill, NSW, Australia), and MRS 2179 (Abcam Biochemicals, Melbourne, Australia). Stock solutions of antagonists were initially made up in distilled water and diluted to working concentrations on the day of the experiment.
5
Pharmacological Modulation of P2Y Receptors
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All chemicals were purchased from Sigma-Aldrich (UK) with the exception of BMSCCR222, BAPTA AM, MRS2578, ARL67156, AR-C66096 and Bordetella pertussis toxin (PTx; Tocris Bioscience, UK), MRS2179, MRS2211 (Abcam Biochemicals, UK), CCL2, TNFα and Fluo-4 AM (Life Technologies, UK), and Calcein AM (Santa Cruz Biotechnology, Germany). THP-1 and HEK 293T cells were procured from the European Collection of Cell Cultures (ECACC), and HUVECs from Caltag Medsystems (UK). Human 1321N1 P2Y6 stable cells were a kind gift from Jens Leipziger (Aarhus University, Denmark). Compounds used did not induce toxicity under assay conditions as determined by a Trypan Blue exclusion assay or lactate dehydrogenase (LDH) release assay.
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!