Feces were collected in Fast Prep lysing Matrix A tubes (MP Biomedicals), resuspended in 1ml of PBS per 100mg fecal material and incubated at 4°C for 20 min. Bacterial suspensions were resuspended in a final volume of 2 ml PBS and incubated at 4°C for 20 min. Samples were homogenized in a FastPrep-24 Tissue homogenizer (MP Biomedicals) for 30s. After homogenization, samples were centrifuged at 50 x g for 15 minutes at 4°C to remove debris and the bacteria-containing supernatant transferred through 70μm filters into a new tube. Bacteria were washed in FACs buffer (PBS, 2% FCS, 5mM EDTA) and pelleted at 8000 x g for 5 min. For flow cytometry, bacterial pellets were resuspended in 100μl FACs buffer containing SYTO 9 green fluorescent nucleic stain (Life Technologies) (10μM), incubated at 4°C for 15 minutes, and subsequently stained with 1μg/ml of an anti-mouse IgA-PE antibody (clone mA-6E1, eBioscience) for 30 min at 4°C. Samples were thoroughly washed and acquired on a BD Fortessa flow cytometer.
Anti mouse iga pe antibody clone ma 6e1
Manufactured by Thermo Fisher Scientific
The Anti-mouse IgA-PE antibody (clone mA-6E1) is a laboratory reagent used for the detection and quantification of mouse IgA antibodies. It is conjugated with the fluorescent dye phycoerythrin (PE), which allows for the visualization and measurement of IgA-positive cells or samples using flow cytometry or other fluorescence-based techniques.
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2 protocols using anti mouse iga pe antibody clone ma 6e1
1
Isolation and Analysis of Gut Bacteria
2
Isolation and Analysis of Gut Bacteria
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Feces were collected in Fast Prep lysing Matrix A tubes (MP Biomedicals), resuspended in 1ml of PBS per 100mg fecal material and incubated at 4°C for 20 min. Bacterial suspensions were resuspended in a final volume of 2 ml PBS and incubated at 4°C for 20 min. Samples were homogenized in a FastPrep-24 Tissue homogenizer (MP Biomedicals) for 30s. After homogenization, samples were centrifuged at 50 x g for 15 minutes at 4°C to remove debris and the bacteria-containing supernatant transferred through 70μm filters into a new tube. Bacteria were washed in FACs buffer (PBS, 2% FCS, 5mM EDTA) and pelleted at 8000 x g for 5 min. For flow cytometry, bacterial pellets were resuspended in 100μl FACs buffer containing SYTO 9 green fluorescent nucleic stain (Life Technologies) (10μM), incubated at 4°C for 15 minutes, and subsequently stained with 1μg/ml of an anti-mouse IgA-PE antibody (clone mA-6E1, eBioscience) for 30 min at 4°C. Samples were thoroughly washed and acquired on a BD Fortessa flow cytometer.
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