实验方案：
体内巨噬细胞清除/耗竭实验
参照既往文献并稍加改进开展巨噬细胞耗竭实验。简述如下：选取 8~12 周龄Prkcd野生型（Prkcd+/+）与敲除型（Prkcd−/−）雌性小鼠，按前述方法原位接种 2.5×105 个 E0771 细胞。将两种基因型小鼠分别随机分组，第 0 天经腹腔注射抗 Ly6C 抗体或大鼠 IgG2a 同型对照抗体（每只小鼠给药 100 μg）；第 1 天依据产品说明书注射氯膦酸盐脂质体或对照空脂质体（每只小鼠给药 200 μl）。抗 Ly6C 抗体 / 大鼠 IgG2a 分别于第 0、4、9 天给药；氯膦酸盐脂质体 / 对照脂质体分别于第 1、5、10 天给药。持续监测肿瘤体积，直至实验第 14 天终止实验。
清除效果：
为探究巨噬细胞中 PKCδ 缺失是否对抑制肿瘤、激活 T 细胞起到关键作用，本研究联合使用抗 Ly6C 单克隆抗体与荷兰Liposoma氯膦酸盐脂质体（CP-005-005），对接种 E0771 肿瘤细胞的Prkcd+/+（野生型）及Prkcd−/−（PKCδ 敲除型）小鼠进行巨噬细胞耗竭处理（图 3A）。
与以往研究结果一致（41-43），我们发现*耗竭巨噬细胞可显著延缓野生型（Prkcd+/+）小鼠的肿瘤生长*（图 3B、D）。与之不同的是，对 Prkcd−/−小鼠进行巨噬细胞耗竭处理后，肿瘤生长并未受到抑制，反而出现明显加快，其肿瘤生长水平与Prkcd+/+ 小鼠相近（图 3C、D）。
论文信息：
论文题目：Protein kinase C delta regulates mononuclear phagocytes and hinders response to immunotherapy in cancer
期刊名称：Science Advances
时间期卷：Vol 7, Issue 51(2023)
在线时间：2023年12月22日
DOI: 10.1126/sciadv.add3231
产品信息：
货号：CP-005-005
规格：5ml+5ml
品牌：Liposoma
产地：荷兰
名称：Clodronate Liposomes&Control Liposomes
办事处：靶点科技
Clodronate Liposomes氯膦酸盐脂质体在敲除小鼠肿瘤模型种清除巨噬细胞。荷兰Liposoma巨噬细胞清除剂ClodronateLiposomes见刊于Science Advances：蛋白激酶 Cδ 调控单核吞噬细胞，并削弱肿瘤的免疫治疗应答。
Liposoma巨噬细胞清除剂Clodronate Liposomes氯膦酸二钠脂质体清除巨噬细胞的材料和方法：
All reagents were obtained from Sigma-Aldrich (St. Louis, MO) unless otherwise noted. Fetal bovine serum (FBS; Gibco, Waltham, MA), 100× l-glutamine, 100× penicillin/streptomycin HyClone (Pittsburgh, PA), and Gibco 100× antibiotic mix were obtained from Thermo Fisher Scientific (Waltham, MA). RPMI 1640, Dulbecco’s modified Eagle’s medium (DMEM), and Matrigel are from Corning (Tewksbury, MA). Mouse recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin- 6 (IL-6), IL-4, macrophage CSF (M-CSF), and FMS-like tyrosine kinase 3 ligand (FLT3L) were obtained from BioLegend (San Diego, CA). OVA was obtained from Thermo Fisher Scientific. Mouse IFN-γ ELISA kit was obtained from R&D Systems (Minneapolis, MN). Mouse CD4+ T cell isolation kit and CD8+ T cell isolation kit were obtained from Miltenyi Biotec (Auburn, CA). Clodronate and control liposomes were obtained from Liposoma (Amsterdam, The Netherlands). In vivo anti-mouse CD40, anti-mouse PD-1, anti-mouse Ly6C monoclonal antibodies, and their controls (rat IgG2a) were all obtained from Bio X Cell (Lebanon, NH). KO-validated PKCδ antibody and phycoerythrin/Cy7 conjugation kit were obtained from Abcam (Cambridge, UK). Flow cytometry antibodies, compensation beads, and reagents are described in table S1 [Tonbo Biosciences Inc. (San Diego, CA), Thermo Fisher, and BioLegend].
巨噬细胞清除材料和方法文献截图：
