Poly(lactic-co-glycolic acid) (PLGA, lactide:glycolide = 50:50, molecular weight [MW] 15,000 Da) was purchased from Jinan Daigang Biomaterial Co., Ltd. (Shandong, China). poly(vinyalcohol) (PVA, MW 25,000 Da) and polyethylenimine (PEI, MW 1800 Da) were purchased from Sigma-Aldrich (St Louis, MO, USA). Paclitaxel (PTX) was purchased from Aladdin Bio-Chem Technology Co., Ltd (Shanghai, China). Cu-tetrakis(4-carboxyphenyl)porphyrin and FITC-labeled PTX were purchased from Ruixi Biotech Co., Ltd (Xi’an, China). tLyP-1 (CGNKRTR) was synthesized by Chinapeptide (Shanghai, China). MCF-7, MCF-7/Taxol and HUVECs were purchased from the MEIXUAN Biological science and technology Co., Ltd (Shanghai, China). GSH assay kit was purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China). LysoTracker Green DND-26 and agarose were purchased from Invitrogen (Thermo Fisher Scientific). Cell Counting Kit-8 (CCK-8) Calcein-AM, and PI were obtained from Dojindo (Japan). P-gp Rabbit pAb was obtained from ABclonal (Wuhan, China). Anti-PD-L1 was purchased from BioXcell (USA). Tumor necrosis factor alpha, interferon gamma and interleukin 6 ELISA kit were purchased from Uscn Life Science, Inc., (China). Anti-CD3+-FITC, anti-CD8+a-APC and anti-CD4+-PC5.5 were purchased from Biolegend (USA). Deionized water obtained from the Millipore system (Direct-Q 5, FRA) was used in all preparations.
Anti cd8a apc
Manufactured by BioLegend
Sourced in United States
Anti-CD8a-APC is a fluorochrome-conjugated antibody used to detect and quantify CD8a-positive cells in flow cytometry applications. It binds specifically to the CD8a cell surface marker, which is expressed on cytotoxic T cells and a subset of natural killer cells.
Automatically generated - may contain errors
Lab products found in correlation
Anti cd4 fitc, by BioLegend (4 mentions)
Anti cd3 fitc, by BioLegend (2 mentions)
Cell strainer, by BD (2 mentions)
Monensin, by Yeasen (2 mentions)
Fixation buffer, by BioLegend (2 mentions)
Anti ifn γ bv 421, by BioLegend (2 mentions)
Anti cd3 pe, by BioLegend (2 mentions)
Anti cd11b pe, by BioLegend (2 mentions)
Agarose, by Thermo Fisher Scientific (1 mentions)
Anti pd l1, by BioXCell (1 mentions)
Lysotracker green dnd 26, by Thermo Fisher Scientific (1 mentions)
Cell counting kit 8 cck 8 calcein am, by Dojindo Laboratories (1 mentions)
Polyethylenimine (pei), by Merck Group (1 mentions)
Gsh assay kit, by Nanjing Jiancheng (1 mentions)
Facsaria cell sorter, by BD (1 mentions)
Intracellular staining buffer, by BioLegend (1 mentions)
Anti cd4 apc cy7, by BioLegend (1 mentions)
Anti cd11b pe cy7, by BioLegend (1 mentions)
Anti mouse cd16 cd32, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
12 protocols using anti cd8a apc
1
Paclitaxel-Loaded PLGA Nanoparticles
2
Thymocyte Subpopulation Isolation in Mice
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Thymus tissues of female BALB/c WT mice were cut into 2-mm pieces, then mechanically disintegrated and passed through cell strainers (BD Bioscience) to obtain the single cell suspensions. Red blood cells of single cell suspensions were lysed. The cells were stained with anti-CD4-FITC (Biolegend) and anti-CD8a-APC (Biolegend) for 30 min on ice, then sorted with a FACS Aria Cell Sorter (BD Bioscience). The thymocyte subpopulations were identified as CD4-CD8- (double negative, DN), CD4+CD8+ (double positive, DP), CD4+ CD8- (CD4+ single positive, CD4+ SP), and CD4- CD8+ (CD8+ single positive CD8+ SP).
3
Functional T Cell Responses in SARS-CoV-2 Vaccinated Animals
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Functional responses of SARS-CoV-2 RBD dodecamer specific CD8+ and CD4+ T cells in vaccinated animals were measured using peptide pools and an intracellular cytokine staining (ICS) assay. Splenocytes were isolated in complete RPMI 1640 medium (HyClone Laboratories Inc., SH30197.02, Grand Island, NY, USA). Cells were incubated overnight with a peptide pool of Delta S proteins (DG peptides) and 2 μmol/L monensin (YEASEN, HB210319, Shanghai, China) was added after 2 h. Cells were harvested and stained with anti-CD4 FITC (Biolegend, 100,405, San Diego, CA, USA) and anti-CD8a APC (Biolegend, 162,305, San Diego, CA, USA), and then fixed and permeabilized in fixation buffer (Biolegend, 420,801, San Diego, CA, USA) and intracellular staining buffer (Biolegend, 421,002, San Diego, CA, USA), followed by anti-interferon γ (IFN-γ) PE/Cy7, anti-interleukin-2 (IL-2) PE, and anti-interleukin-4 (IL-4) PerCP/Cy5.5 staining, and were analyzed and quantified by flow cytometry (NovoCyteTM, Eisen Bioscience, USA).
4
Multiparametric Immune Profiling of Lung Metastases
5
Multiparameter Immune Cell Profiling
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Anti–CD3ε-PerCP (100326), anti–CD8a-AF700 (100730), anti–CD8a-APC (100712), anti–PD-1-PE/DZ594 (109116), anti–Ki67-FITC (652410), anti–TCF1-PE (655208), anti–IFN-γ-BV421 (505830), anti–IFN-γ-PerCp/Cy5.5 (505822), anti–TNF-α-APC/Cy7 (506344), anti–TIM-3-PE/Cy7 (119716), anti–CD45-BV510 (103138), anti–Ly108 (Slamf6)-PE (134606), anti–PD-1-FITC (135214), anti–CD45.1-PE (110708), anti–CD45.2-AF488 (109816), purified anti-mouse CD3ε (100340), and purified anti-mouse CD28 (102116) were obtained from Biolegend. Anti–CD8-FITC (D271-4) and tetramer-SIINFEKL-APC (TS-5001-2c) were purchased from MBL. Cell Stimulation Cocktail Plus Protein Transport Inhibitors and FOXP3/TF Staining Buffer Set were bought from eBioscience. Naive CD8a+ T Cell Isolation and Tumor Dissociation Kits were obtained from Miltenyi Biotec.
6
Suppression of PBMC Proliferation by MDSC
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MDSC were generated from myeloid cells of the PBMC fraction as described above and isolated from cell cultures by magnetic bead cell sorting for CD33 (Miltenyi Biotec). Responder-PBMC were obtained from healthy volunteers' heparinized blood and stained with CFSE (Life Technologies) according to the manufacturer's protocol. CFSE-labeled PBMC were stimulated with 100 U/ml IL-2 (R&D Systems) and 1 μg/ml OKT3 (Janssen-Cilag). Both MDSC and CFSE-labeled PBMC were added to RPMI 1640 medium supplemented with 10% human serum, 2 mM L-glutamine, 100 IU/ml penicillin and 100 mg/ml streptomycin. In a 96-well round bottom plate (Greiner Bio-One), either 10,000/30,000 MDSC or, as a control supplemented medium only, were added to 60,000 PBMC per well. Cells were incubated in a humidified atmosphere at 37°C and 5% CO2. On day 4 cells were harvested and stained with anti-CD8a-APC, anti-CD4-PE antibodies (BioLegend), and propidium iodide (BD). PI positive cells were excluded in flow cytometry. CFSE signals of CD4+ and CD8+ PBMC were analyzed.
7
Immune Cell Profiling in Tumor Microenvironment
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The mice were euthanized 24 hours after the last PET imaging timepoint. To study the immune cells in tumors, tumors from different groups were collected by surgery, then homogenized into single-cell suspensions according to the well-established procedure. To analyze the effector T cells (CD45+, CD3+, CD8+), tumor cells were stained with anti-CD45-FITC (Biolegend, USA), anti-CD3-PE (Biolegend, USA) and anti-CD8a-APC (Biolegend, USA) antibodies according to the standard protocol. TAM cells were classified into M1 TAM (CD11b+, F4/80+, CD86+) and M2 TAM cells (CD11b+, F4/80+, CD206+). For analysis of TAM cells, tumor cells were stained with anti-CD11b-PE (Biolegend, USA), anti-F4/80-FITC (Biolegend, USA), anti-CD86-Percp/Cyanine5.5 (Biolegend, USA), anti-CD206-APC (Biolegend, USA) antibodies and examined using flow cytometry. For DCs maturation (CD11c+, CD80+, CD86+) analysis, spleens collected from mice after different treatments were homogenized into single-cell suspensions and stained with anti-CD11c-FITC (Biolegend, USA), anti-CD80-APC (Biolegend, USA), anti-CD86-Percp/Cyanine5.5 antibodies for flow cytometry examination. All these antibodies used in ex vivo flow cytometry experiments were diluted 150 times.
8
Tumor-Infiltrating T-Cell Analysis
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At the time of sacrifice, the tumors and spleens were harvested from the mice. Single-cell suspensions of splenocytes were stained with anti-CD3-FITC (cat. no. 11-0032-82; eBioscience, San Diego, USA), anti-CD4-PerCP/Cy5.5 (cat. no. 100540; BioLegend) and anti-CD8a-APC (cat. no. 17-0081-83; eBioscience) at 4 °C for 20 min. The single-cell suspensions of tumors were stained with anti-CD45-R-PE (cat. no. 147711; BioLegend), anti-CD3-FITC, anti-CD8a-APC and anti-IFN-γ-Bv421 (cat. no. 505830; BioLegend) at 4 °C for 20 min. The cells were analyzed by an Attune NxT Flow Cytometer.
9
Cytokine Production Assay for E7 Peptide
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The cells were incubated at a concentration of 3x106 cells/well for 6 h at 37 ºC and 5% CO2 with 10 µg/mL Brefeldin A (GolgiPlug; BD Biosciences), 5 ng/mL IL-2 (Thermo Fisher Scientific), and 300 ng/well of E7 peptide16 (amino acids 49-57; RAHYNIVTF; GenScript). After incubation, cells were stained for 30 min at 4 ºC with anti-CD8a-APC (BioLegend), anti-CD44-FITC (BioLegend), anti-CD62L-BV421 (BioLegend), anti-KLRG1-PE (BioLegend), and anti-CD127-PECy7 (BD Biosciences) antibodies. After fixation/permeabilization with the Cytofix/Cytoperm solution (BD Biosciences) for 10 min at 4 ºC, cells were stained with anti-IFN-γ antibody conjugated to Alexa700 (BD Biosciences) for 30 min at 4 ºC. The cells were then resuspended in PBS and examined by flow cytometry using the LSR Fortessa device (BD Biosciences).
10
Multiparametric Flow Cytometry of Immune Cells
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Lung and thymus tissues were cut into 2-mm pieces, then mechanically disintegrated and passed through cell strainers (BD Bioscience) to obtain the single cell suspensions. Red blood cells from peripheral blood were directly lysed to obtain single cell suspensions. First, Fixable Viability Stain 450 (562247, BD Bioscience) was performed for live-dead marker in the absence of other antibodies. The cells were incubated with anti-CD16/32 (101301, Biolegend) for blocking Fc receptor. Dividing cells were stained with anti-CD3-PE (Biolegend), anti-CD4-FITC (Biolegend) and anti-CD8a-APC (Biolegend) for 30 min on ice. To determine intracellular cytokine levels, cells were further fixed, permeabilized, and stained with anti-IFN-γ-AlexaFluor 700 (BD Bioscience) and anti-IL-4-APC (BD Bioscience). Data were acquired with LSR Fortessa flow cytometer (BD Bioscience), and analyzed with the Tree Star Flowjo software.
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