Adoptive Transfer of Spleen Lymphocytes and Immunoglobulins for Antitumor Efficacy

Preparation of spleen lymphocytes was performed as previously reported44 (link). Briefly, the immunized or control mice were killed on day 7 after the third immunization. Single-cell suspensions were obtained from spleens through a 70-μm nylon mesh filter (BD Biosciences), and lymphocytes were enriched by lymphocyte separation medium (Dakewe Biotech Company) according to the manufacturer’s instructions. To assess the efficacy of T cells in antitumor in vivo, spleen lymphocytes (1 × 10⁷) from donor mice on day 7 after the third immunization were adoptively transferred intravenously into recipient mice one day before and three days after the tumor inoculation (3 × 10⁶). Immunoglobulins were purified from the pooled sera derived from the immunized or control mice by affinity chromatography (CM Affi-gel Blue Gel Kit; Bio-Rad). To assess the efficacy of Immunoglobulins in antitumor activity in vivo, the purified Immunoglobulins (50 mg/kg) were adoptively transferred intravenously one day before mice were challenged with 3 × 10⁶ tumor cells and then treated twice per week for 3 weeks.