Macrophages were obtained from mouse lungs (n=5) after MRI protocol at 2 hours post-antibody-conjugated SPIO nanoparticle administration. They were prepared for flow cytometry as per the protocol described elsewhere.21 (link) Briefly, the dissected lungs were held in complete RPMI (Roswell Park Memorial Institute) medium supplemented with 25 mM HEPES (2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid). Lungs were minced in complete medium, and the filtrates were centrifuged (1,500 rpm for 5 minutes). The number of cells in the supernatant was determined, and then the solutions were passed through magnetic activated cell sorter (MACS) microbeads and through MACS LD or MS columns, as per the manufacturer’s protocol (Miltenyi Biotec). The eluted unbound and bound cell fractions were first separated, and after a repeat cell count, they were tested for different markers by flow cytometry. The cells were then washed, fixed, and permeabilized with Cytofix/Cytoperm™ (BD Biosciences). They were stained with either Alexa Fluor® 488-labeled anti-CD86 or FITC-labeled anti-CD206 anti-mouse antibodies and analyzed with the BD™ LSR II flow cytometer.
Macs ld or ms columns
Manufactured by Miltenyi Biotec
Sourced in Germany
The MACS LD or MS columns from Miltenyi Biotec are laboratory equipment used for cell separation and enrichment. They are designed to work with the company's MACS cell separation technology, allowing for the efficient isolation and purification of specific cell populations from complex samples.
Automatically generated - may contain errors
Lab products found in correlation
Cytofix cytoperm, by BD (2 mentions)
Macs microbead, by Miltenyi Biotec (2 mentions)
Lsr 2 flow cytometer, by BD (1 mentions)
Alexa fluor 647 labeled anti cd197, by Bio-Rad (1 mentions)
Fitc labeled anti cd206, by Bio-Rad (1 mentions)
Cellquest software, by BD (1 mentions)
Facscalibur flow cytometry system, by BD (1 mentions)
2 protocols using macs ld or ms columns
1
Isolation and Characterization of Macrophages from Mouse Lungs
2
Isolation and Analysis of Lung Macrophages
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Macrophages were obtained from mouse lungs after sacrifice and prepared for flow cytometry as per the protocol described elsewhere [23] (link). Briefly, the dissected lungs were held in complete RPMI medium supplemented with 25 mM HEPES. Lungs were minced in complete medium and the filtrates were centrifuged (1500 rpm for 5 min). The number of cells in the supernatant were determined and then the solutions passed through magnetic activated cell sorter (MACS) microbeads and through MACS LD or MS columns, as per the manufacturer’s protocol (Miltenyi Biotec, Germany). The eluted unbound and bound cell fractions were first separated, and after a repeat cell count, they were tested for different markers by flow cytometry. The cells were then washed, fixed and permeabilized with Cytofix/Cytoperm (BD Biosciences, NJ, USA), stained with conjugated anti-mouse antibodies as follows: Alexa Fluor 488-labeled anti-CD86, Alexa Fluor 647-labeled anti-CD197, RPE-labeled anti-CD23, and FITC-labeled anti-CD206 (AbD Serotec, UK), and analyzed with BD LSR II flow cytometer using DIVA Software. Data were collected (10,000 events each time) with a FACSCalibur flow cytometry system and analyzed with the CellQuest Software (BD Biosciences, NJ, USA).
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