Splenocytes were isolated from OT-I mice and labeled with 5μM CFSE for 9 min at 37°C. Reactions were quenched with FBS, and CD8+ T cells were isolated with a negative selection magnetic microbeads kit (Miltenyi Biotec), according to the manufacturer’s protocol. Splenocytes were harvested from wild-type mice and irradiated for 15 minutes at 3000 rads. T cells were depleted using magnetic microbeads (Miltenyi Biotec) and the remaining splenocytes were pulsed with 1μM OT-I peptide (SIINFEKL). Cells were cultured together for 3 days and analyzed on an Accuri C6 (BD Biosciences) flow cytometer with FlowJo software (Treestar).
Negative selection magnetic microbeads kit
Manufactured by Miltenyi Biotec
The Negative selection magnetic microbeads kit is a laboratory tool used for the isolation and purification of specific cell populations from a heterogeneous sample. The kit utilizes magnetic beads coated with antibodies that bind to and remove unwanted cells, allowing the desired cells to be collected in a purified form.
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2 protocols using negative selection magnetic microbeads kit
1
Antigen-specific CD8+ T cell assay
2
Splenocyte Isolation and T Cell Activation
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Splenocytes were isolated from OT-I mice and labeled with 5μM CFSE for 9 min at 37 °C. Reactions were quenched with FBS, and CD8+ T cells were isolated with a negative selection magnetic microbeads kit (Miltenyi Biotec), according to the manufacturer’s protocol. Splenocytes were harvested from wild-type mice and irradiated for 15 min at 3000 rads. T cells were depleted using magnetic microbeads (Miltenyi Biotec) and pulsed with 1μM OT-I peptide (SIINFEKL). Cells were cultured together for 3 days and analyzed on an Accuri C6 (BD Biosciences) with FlowJo software (Treestar).
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