實驗方案：

體內(nèi)巨噬細胞清除/耗竭實驗

參照既往文獻并稍加改進開展巨噬細胞耗竭實驗。簡述如下：選取 8~12 周齡Prkcd野生型（Prkcd^+/+）與敲除型（Prkcd^?/?）雌性小鼠，按前述方法原位接種 2.5×105 個 E0771 細胞。將兩種基因型小鼠分別隨機分組，第 0 天經(jīng)腹腔注射抗 Ly6C 抗體或大鼠 IgG2a 同型對照抗體（每只小鼠給藥 100 μg）；第 1 天依據(jù)產(chǎn)品說明書注射氯膦酸鹽脂質(zhì)體或?qū)φ湛罩|(zhì)體（每只小鼠給藥 200 μl）?？?Ly6C 抗體 / 大鼠 IgG2a 分別于第 0、4、9 天給藥；氯膦酸鹽脂質(zhì)體 / 對照脂質(zhì)體分別于第 1、5、10 天給藥。持續(xù)監(jiān)測腫瘤體積，直至實驗第 14 天終止實驗。

清除效果：

為探究巨噬細胞中 PKCδ 缺失是否對抑制腫瘤、激活 T 細胞起到關(guān)鍵作用，本研究聯(lián)合使用抗 Ly6C 單克隆抗體與荷蘭Liposoma氯膦酸鹽脂質(zhì)體（CP-005-005），對接種 E0771 腫瘤細胞的Prkcd+/+（野生型）及Prkcd?/?（PKCδ 敲除型）小鼠進行巨噬細胞耗竭處理（圖 3A）。

與以往研究結(jié)果一致（41-43），我們發(fā)現(xiàn)*耗竭巨噬細胞可顯著延緩野生型（Prkcd+/+）小鼠的腫瘤生長*（圖 3B、D）。與之不同的是，對 Prkcd?/?小鼠進行巨噬細胞耗竭處理后，腫瘤生長并未受到抑制，反而出現(xiàn)明顯加快，其腫瘤生長水平與Prkcd+/+ 小鼠相近（圖 3C、D）。

論文信息：

論文題目：Protein kinase C delta regulates mononuclear phagocytes and hinders response to immunotherapy in cancer

期刊名稱：Science Advances

時間期卷：Vol 7, Issue 51(2023)

在線時間：2023年12月22日

DOI: 10.1126/sciadv.add3231

產(chǎn)品信息：

貨號：CP-005-005

規(guī)格：5ml+5ml

品牌：Liposoma

產(chǎn)地：荷蘭

名稱：Clodronate Liposomes&Control Liposomes

辦事處：靶點科技

Clodronate Liposomes氯膦酸鹽脂質(zhì)體在敲除小鼠腫瘤模型種清除巨噬細胞。荷蘭Liposoma巨噬細胞清除劑ClodronateLiposomes見刊于Science Advances：蛋白激酶 Cδ 調(diào)控單核吞噬細胞，并削弱腫瘤的免疫治療應(yīng)答。

Liposoma巨噬細胞清除劑Clodronate Liposomes氯膦酸二鈉脂質(zhì)體清除巨噬細胞的材料和方法：

Macrophage depletion

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].

巨噬細胞清除材料和方法文獻截圖：