Immediately after photic injury was induced, the mice received subcutaneous injections of either the CCR1-specific antagonist BX471 (50 mg/kg body weight; Tocris, Bristol, UK) or vehicle (40% cyclodextrin in saline) every 12 hr for 5 days. For injection, BX471 was dissolved at a final concentration of 10 mg/ml in saline containing 40% (wt/vol) cyclodextrin (Sigma-Aldrich); the solution was mixed thoroughly and dissolved overnight at 4°C, after which the pH was adjusted to 4.5 with NaOH, and the solution was filtered through a 0.45 µm filter.
Bx471
Manufactured by Bio-Techne
Sourced in United Kingdom
The BX471 is a piece of laboratory equipment manufactured by Bio-Techne. It is designed to perform specific tasks within a research or testing environment. The core function of the BX471 is to support various laboratory processes, but a detailed description cannot be provided while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
Lab products found in correlation
Bx471, by Merck Group (1 mentions)
Px 478, by Selleck Chemicals (1 mentions)
Sb297006, by Bio-Techne (1 mentions)
Dapta, by Bio-Techne (1 mentions)
Ly294002, by Selleck Chemicals (1 mentions)
Sb328437, by Bio-Techne (1 mentions)
Rs504393, by Bio-Techne (1 mentions)
Nc sirna, by GenePharma (1 mentions)
3 well slides, by Ibidi (1 mentions)
Culture insert 2 well, by Ibidi (1 mentions)
Protein low bind tubes, by Eppendorf (1 mentions)
4 protocols using bx471
1
Subcutaneous CCR1 Antagonist Treatment Post-Photic Injury
2
Conditioned Media Collection and Inhibition
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For the following series of experiments, cells were first cultured in RPMI‐1640 complete media (5% FBS) in 6‐well plates. After reaching confluence, the cells were washed with PBS and cultured in serum‐free media (2 mL/well) for 24 hours. Cell supernatants were collected as the CM and stored at −80°C until use. For collecting CM in chemical inhibition experiments, the PI3K inhibitor LY294002 (Selleckchem, Houston, TX, USA), the HIF‐1α inhibitor PX478 (Selleckchem), the CCR1 antagonist BX471 (Tocris, Bristol, UK), the CCR3 antagonist SB297006 (Tocris), or the CCR5 antagonist DAPTA (Tocris) was used to pretreat cells for 6 hours. The cells were then washed and cultured in serum‐free media for collecting CM.
3
Trigeminal Ganglion Injection Protocol
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The recombinant mouse CCL7 was purchased from Sino Biological (Beijing, China). Ccl7 siRNA and negative control siRNA (NC siRNA) were designed by Gene Pharma (Suzhou, China). The selective CCR1 antagonist BX471, CCR2 antagonist RS504393, and CCR3 antagonist SB328437 were purchased from Tocris (Bristol, UK). For Intra-TG injection, the animals were anesthetized with isoflurane and drugs were injected with a 30 G needle from the infraorbital foramen to the foramen rotundum. The tip of the needle terminated at the medial part of the TG, and the siRNA or antagonists (5 μL) were slowly delivered.
4
Generating Conditioned Media for Cell Migration Assays
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To generate conditioned media for migration assays, cells were cultured in growth medium without FBS. After two days, the medium was aspirated, centrifuged at 1000× g, and stored in Protein LowBind® tubes (Eppendorf, Hamburg, Germany) at −20 °C until use. For migration assays, 3.5 × 105 cells/mL were cultured in removable 2-well culture inserts inside 3-well slides (both Ibidi, Gräfeling, Germany). After an adherence period of 24 h in the growth medium, cells were washed twice with serum-free medium and then cultured for 24 h in serum-free medium for FBS starvation. The 2-well chambers were removed and then washed with serum-free medium, and the experimental conditions were applied. For migration assays using MCF-7, 6 × 105 cells/mL were seeded per chamber. After an adherence period of 24 h in growth medium, cells were washed twice with serum-free medium, and conditions were applied. CCR1-antagonist BX471 (Tocris Bioscience, Bristol, UK) was used at a concentration of 30 µM, while CCR5 antagonist TAK-779 (Merck, Darmstadt, Germany) was used at a concentration of 1 µM [30 (link)]. Gap closure was documented after 0 h and 48 h and the percentage of area covered was assessed using FastTrackAI (MetaVi Labs, Austin, TX, USA).
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