Separation buffer: Prepare a solution containing phosphate-buffered saline (PBS), pH 7.2, and 0.5% bovine serum albumin (BSA), and 2 mM EDTA by diluting MACS® BSA Stock Solution (# 130-091-376) 1:20 with autoMACS® Rinsing Solution (#130-091-222). Keep buffer cold (2−8 °C). Degas buffer before use, as air bubbles could block the column. Always use freshly prepared buffer.
▲Note: EDTA can be replaced by other supplements, such as anticoagulant citrate dextrose formula-A (ACD-A), or citrate phosphate dextrose (CPD). BSA can be replaced by other proteins, such as mouse serum albumin, mouse serum, or fetal bovine serum (FBS). Buffers or media containing Ca2+ or Mg2+ are not recommended for use.
REAlease Bead Release buffer: Prepare a 1:50 dilution of REAlease Bead Release Reagent (50×), e.g., for 1 mL add 20 μL of REAlease Bead Release Reagent to 980 μL of separation buffer.
▲Note: Use buffer freshly prepared at the same day. Store at room temperature.
▲Note: Prepare 1 mL per MS Column and 5 mL per LS Column.
Carefully create a single-cell suspension from a solid human tumor using the gentleMACS™ Octo Dissociator with Heaters in combination with the Tumor Dissociation Kit, human. For detailed instructions, follow the protocol of the kit data sheet.
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