Enrichment of bead- and label-free human tumor-infiltrating leukocytes (TILs) from human tumor tissue

• Tumor Dissociation Kit, human (# 130-095-929)
• gentleMACS™ Octo Dissociator with Heaters (# 130-096-427)
• gentleMACS C Tubes (# 130-093-237)
• Culture media
• MACS® SmartStrainers (70 µm) (# 130-098-462)
• MACSmix™ Tube Rotator (# 130-090-753) in combination with an incubator at 37 °C
• (Optional) Red Blood Cell Lysis Solution (10×) (# 130-094-183)
• (Optional) MACS Tissue Storage Solution (# 130-100-008)

• Select your marker of interest among the following:
  REAlease® CD45 (TIL) MicroBead Kit, human (# 130-121-563)
  REAlease CD3 (TIL) MicroBead Kit, human (# 130-121-562)
  REAlease CD4 (TIL) MicroBead Kit, human (# 130-121-559)
  REAlease CD8 (TIL) MicroBead Kit, human (# 130-121-560)
  REAlease CD4/CD8 (TIL) MicroBead Kit, human (# 130-121-561)
• QuadroMACS™ Starting Kit (LS) (# 130-091-051)
• 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.
• (Optional) Pre-Separation Filters (30 µm) (# 130-041-407) to remove cell clumps
• (Optional) MACS® SmartStrainers (30 µm) (# 130-098-458) to remove cell clumps
• (Optional) Fluorochrome-conjugated recombinant antibodies for flow cytometry analysis, e.g., CD45 Antibody, anti-human, VioBlue®. Learn more about our antibodies for flow cytometry.
  ▲Note: Due to expression of Fcγ receptors on tumor-infiltrating leukocytes, REAfinity™ antibodies are recommended.
• (Optional) Propidium Iodide Solution (# 130-093-233), DAPI Staining Solution (# 130-111-570), 7-AAD Staining Solution (# 130-111-568), or Viobility™ Fixable Dyes (# 130-109-812, # 130-109-814, # 130-109-816) for flow cytometric exclusion of dead cells.
• (Optional) Dead Cell Removal Kit (# 130-090-101) for the depletion of dead cells.

Unless otherwise specifically indicated, Miltenyi Biotec products and services are for research use only and not for therapeutic or diagnostic use. 

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