Pe conjugated anti cd3

Manufactured by BD

Sourced in United States

PE-conjugated anti-CD3 is a monoclonal antibody that binds to the CD3 complex on the surface of T cells. The PE (phycoerythrin) fluorescent dye is conjugated to the antibody, allowing for the detection and identification of CD3-positive T cells in flow cytometry applications.

Automatically generated - may contain errors

Lab products found in correlation

12 protocols using pe conjugated anti cd3

Quantifying CAR T-cell Transduction Efficacy

Check if the same lab product or an alternative is used in the 5 most similar protocols

To validate the CAR transduction efficacy and the changes of CAR T-cells in the blood after injection, we performed flow cytometry assay.
For CAR transduction efficacy, CAR-T cells (1 × 10⁶) were suspended in 100 μL Dulbecco’s PBS (DPBS; Thermo Fisher) and incubated with PE-conjugated anti-CD3 (BD, Biosciences) and FITC-conjugated anti-CAR (Immunochina Pharmaceuticals) for 30 min. After washing with DPBS twice, the cells were evaluated with FlowJo software (FlowJo 7.6.1).
For CAR-T cell detection, red blood cells were removed using an RBC lysing buffer (Sigma Aldrich, MO) for 5 min, followed by washing and re-suspension in 1× HBSS containing 1% FBS. The separated blood cells were stained with PE-conjugated anti-CD3 (BD, Biosciences) and FITC-conjugated anti-CAR (Immunochina Pharmaceuticals) in 4 °C for 30 min and followed by washing with DPBS containing 1% FBS. Cells were analyzed using BD FACS Callibur cytometry (BD Biosciences). The results were evaluated by FlowJo software.

Ying Z., He T., Wang X., Zheng W., Lin N., Tu M., Xie Y., Ping L., Zhang C., Liu W., Deng L., Wu M., Feng F., Leng X., Du T., Qi F., Hu X., Ding Y., Lu X.A., Song Y, & Zhu J. (2021). Distribution of chimeric antigen receptor-modified T cells against CD19 in B-cell malignancies. BMC Cancer, 21, 198.

+ Open protocol

+ Expand

Multiparameter Flow Cytometry Immunophenotyping

Check if the same lab product or an alternative is used in the 5 most similar protocols

This study used the following mAbs and reagents: Allophycocyanin (APC)-Cy7-conjugated anti-CD3, APC-conjugated anti-CD3, APC-conjugated anti-CD69, and fluorescein isothiocyanate (FITC)-conjugated anti-CD45, FITC-conjugated anti-CD56, FITC-conjugated anti-CD107a, FITC-conjugated anti-IFN-γ, phycoerythrin (PE)-conjugated anti-CD3, PE-conjugated anti-CD56, PE-conjugated anti-CD69, PE-Cy7-conjugated anti-TNF-α, PerCP-conjugated anti-CD3, PerCP-conjugated anti-CD45, FITC-conjugated mouse IgG isotype and PE-conjugated mouse IgG isotype control (all from Becton Dickinson, San Diego, CA). Cells were stained with combinations of appropriate mAb for 20 minutes at 4°C. Stained cells were analyzed on a Navios flow cytometer using Kaluza software (version 1.1; Beckman Coulter, Brea, CA).

Kang S.J., Jin H.M., Cho Y.N., Kim S.E., Kim U.J., Park K.H., Jang H.C., Jung S.I., Kee S.J, & Park Y.W. (2017). Increased level and interferon-γ production of circulating natural killer cells in patients with scrub typhus. PLoS Neglected Tropical Diseases, 11(7), e0005815.

+ Open protocol

+ Expand

Immunophenotyping of T Cell Subsets

Check if the same lab product or an alternative is used in the 5 most similar protocols

The following mAbs and reagents were used in this study: Allophycocyanin (APC)-Cy7-conjugated anti-CD3, phycoerythrin (PE)-Cy5-conjugated anti-CD161 and fluorescein isothiocyanate (FITC)-conjugated anti-TCR γδ, FITC-conjugated anti-CD3, FITC-conjugated anti-IFN-γ, FITC-conjugated annexin V, PE-conjugated anti-CD3, PE-conjugated anti-IL-17, PE-Cy7-conjugated anti-TNF-α, PE-conjugated anti-CD69, FITC-conjugated mouse IgG isotype, PE-conjugated mouse IgG isotype and PE-Cy7-conjugated mouse IgG isotype control (all from Becton Dickinson, San Diego, CA); PE-conjugated anti-programmed death-1 (anti-PD-1; eBioscience, San Diego, CA) and APC-conjugated anti-TCR Vα7.2 (BioLegend, San Diego, CA). Cells were stained with combinations of appropriate mAbs for 20 minutes at 4°C. Stained cells were analyzed on a Navios flow cytometer using Kaluza software (version 1.1; Beckman Coulter, Brea, CA).

Kang S.J., Jin H.M., Won E.J., Cho Y.N., Jung H.J., Kwon Y.S., Kee H.J., Ju J.K., Kim J.C., Kim U.J., Jang H.C., Jung S.I., Kee S.J, & Park Y.W. (2016). Activation, Impaired Tumor Necrosis Factor-α Production, and Deficiency of Circulating Mucosal-Associated Invariant T Cells in Patients with Scrub Typhus. PLoS Neglected Tropical Diseases, 10(7), e0004832.

+ Open protocol

+ Expand

MAIT Cell Granzyme and Perforin Detection

Cited 2 times

Check if the same lab product or an alternative is used in the 5 most similar protocols

Granzyme and perforin expression in MAIT cells was detected by intracellular cytokine flow cytometry as previously described [14 (link), 44 (link), 53 (link)]. Briefly, freshly isolated PBMCs (1 × 10⁶/well) were incubated for 12 hours in the presence of PMA (100 ng/ml) and IM (1 μM) or for 24 hours with E. coli-infected THP-1 cells (1 × 10⁶/well; American Type Culture Collection). For intracellular staining, 10 μL of brefeldin A (GolgiPlug; BD Biosciences, San Diego, CA) was added, and the final concentrations of brefeldin A were 10 μg/mL. After incubation for an additional 4 hours, cells were stained with PE-conjugated anti-CD3, PE-Cy5-conjugated anti-CD161 and APC-conjugated anti-TCR Vα7.2 mAb for 20 minutes at 4°C, fixed in 4% paraformaldehyde for 15 minutes at room temperature, and permeabilized with Perm/Wash solution (BD Biosciences) for 10 minutes. Cells were then stained with FITC-conjugated anti-Granzyme and FITC-conjugated anti-perforin mAbs for 30 minutes at 4°C and analyzed by flow cytometry.

Won E.J., Ju J.K., Cho Y.N., Jin H.M., Park K.J., Kim T.J., Kwon Y.S., Kee H.J., Kim J.C., Kee S.J, & Park Y.W. (2016). Clinical relevance of circulating mucosal-associated invariant T cell levels and their anti-cancer activity in patients with mucosal-associated cancer. Oncotarget, 7(46), 76274-76290.

+ Open protocol

+ Expand

MAIT Cell Activation and Cytokine Assay

Check if the same lab product or an alternative is used in the 5 most similar protocols

Jurkat or SKW3 cells (10⁵) transduced with genes encoding MAIT TCRα- and TCRβ-chain genes were tested for activation by co-incubation with compounds (synthetic rRL-6-CH₂OH or 5-A-RU, synthesized as described previously; Corbett et al., 2014 (link)) and C1R.MR1 cells (10⁵) for 16 h. For inhibition, 6-FP, Ac-6-FP, or control pterins (Schircks Laboratories) were added to C1R.MR1 cells for 1 h before addition of Jurkat.MAIT cells and 0.02 µM synthetic rRL-6-CH₂OH. Cells were subsequently stained with PE-conjugated anti-CD3 (BD), and APC-conjugated anti-CD69 (BD) mAbs for 30 min on ice, before analysis of CD69 surface expression by flow cytometry. Activation of Jurkat.MAIT or SKW.MAIT cells was measured by an increase in surface CD69 expression.
Jurkat76 cells (10⁵) transduced with genes encoding the WT original (clone A-F7) MAIT TCR α- and β-chain genes were co-incubated with compounds (6-FP, Ac-6-FP; 5-OP-RU; Schircks Laboratories) and C1R.MR1 cells (10⁵) in 200 µl media for 21 h. IL-2 production was measured as a mean of Jurkat76.MAIT activation in ELISA (BD OptEIA kit) using 100 µl of supernatant, frozen/thawed to kill cells. In brief, IL-2 was assayed with biotinylated anti–IL-2 mAb and o-Phenylenediamine dihydrochloride (OPD; Sigma-Aldrich) substrate conversion by HRP-Streptavidin detected at 492 nm emission.

Eckle S.B., Birkinshaw R.W., Kostenko L., Corbett A.J., McWilliam H.E., Reantragoon R., Chen Z., Gherardin N.A., Beddoe T., Liu L., Patel O., Meehan B., Fairlie D.P., Villadangos J.A., Godfrey D.I., Kjer-Nielsen L., McCluskey J, & Rossjohn J. (2014). A molecular basis underpinning the T cell receptor heterogeneity of mucosal-associated invariant T cells. The Journal of Experimental Medicine, 211(8), 1585-1600.

+ Open protocol

+ Expand

Flow cytometric analysis of GSN and CD3

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cells (1×10⁶ cells/well) were collected by centrifugation (800 × g, 5 min). After the supernatant was decanted, the cells were resuspended in PBS with 1% bovine serum albumin, and incubated with fluorescence-labeled antibodies (anti-GSN [#12953, Cell Signaling Technology, Inc.] with phytoerythrin [PE] Cy5-conjugated anti-rabbit, and PE-conjugated anti-CD3 [BD PharMingen, San Diego, CA, USA]) on ice for 30 minutes. Cells were washed twice with PBS containing 0.1% bovine serum albumin before brief fixation with 1% paraformaldehyde on ice. The number of antibody-labeled cells was then determined by flow cytometry (Beckman Coulter Cytomics^TM FC500), and data were analyzed using machine-embedded CXP software [15 (link)].

Chen C.C., Chiou S.H., Yang C.L., Chow K.C., Lin T.Y., Chang H.W., You W.C., Huang H.W., Chen C.M., Chen N.C., Chou F.P, & Chou M.C. (2017). Secreted gelsolin desensitizes and induces apoptosis of infiltrated lymphocytes in prostate cancer. Oncotarget, 8(44), 77152-77167.

+ Open protocol

+ Expand

Flow Cytometric Analysis of Immune Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

For flow cytometry analysis, cell suspensions were incubated in FACS staining buffer (PBS containing 1% BSA) and subsequently stained for 30 min at 4 °C with an optimized concentration of antibodies (BD Bioscience, Franklin Lakes, NJ, USA): PE-conjugated anti-CD3, PerCPCy5.5-conjugated anti-CD8, PE Cy7-conjugated anti-CD4, PerCPCy5.5-conjugated anti-CD45, APC-conjugated anti-CD11c and PE-conjugated anti-SiglecF to determine cell types in the BAL. Cells were acquired on an LSRII flow cytometer (BD Bioscience) and the data were analyzed using the FlowJo software (v 7.6.5; Ashland, OR, USA). Based on surface marker expression, six different cell types were identified: CD45⁺ (total leukocytes), CD45⁺SiglecF⁺CD11c^low (eosinophils), CD45⁺SiglecF⁺CD11c^high (alveolar macrophages), CD45⁺CD3⁺ (total T cells), CD4 T cells (CD45⁺CD3⁺CD4⁺) and CD8 T cells (CD45⁺CD3⁺CD8⁺).

Wolf S., Johnson S., Perwitasari O., Mahalingam S, & Tripp R.A. (2017). Targeting the pro-inflammatory factor CCL2 (MCP-1) with Bindarit for influenza A (H7N9) treatment. Clinical & Translational Immunology, 6(3), e135-.

+ Open protocol

+ Expand

Multiparametric Flow Cytometry Analysis

Check if the same lab product or an alternative is used in the 5 most similar protocols

Splenocytes from immunized animals were plated at 1 × 10⁶ cells/well in 24-well flat-bottom plate and stimulated with 5 µg of rEF1-α for 12 h at 37°C. Brefeldin A (10 µg/ml) was added to the cultures 2 h before harvesting. The cells were washed in PBS containing 0.1% NaN₃/1% FCS and stained with PE-conjugated anti-CD3 and either PerCP5.5-conjugated anti-CD4 or APC-Cy7-CD8 mAb (BD Pharmingen) for 30 min at 4°C. The cells were then permeabilized with 1× BD Perm2 reagent for 10 min at 4°C, followed by washing with the above-described buffer containing 0.1% saponin and the cells were fixed using cytoperm kit. Then the intracellular cytokines were stained with APC-conjugated anti-IFN-γ, FITC-conjugated anti-IL2 and PE-Cy7-conjugated anti-TNF-α for 30 min at 4°C. After proper washing, cells were analyzed on BD LSR-Fortessa flow cytometer on at least 100,000 events and analyses were carried out by FlowJo software as described previously (33 (link)). For memory markers, the splenocytes apart from CD3-PE, CD4-PE-Cy7, and CD8-APC-Cy7, surface markers were also stained with APC-conjugated anti-CD62L and PerCP5.5-conjugated anti-CD44. After fixation the cells were analyzed similarly as described above.

Sabur A., Bhowmick S., Chhajer R., Ejazi S.A., Didwania N., Asad M., Bhattacharyya A., Sinha U, & Ali N. (2018). Liposomal Elongation Factor-1α Triggers Effector CD4 and CD8 T Cells for Induction of Long-Lasting Protective Immunity against Visceral Leishmaniasis. Frontiers in Immunology, 9, 18.

+ Open protocol

+ Expand

Flow Cytometric Analysis of GD2, PD-L1, and PD-1 Expression

Check if the same lab product or an alternative is used in the 5 most similar protocols

The surface expression of GD2 was analyzed by flow cytometry with PE-conjugated anti-human GD2 mAb clone 14.G2a (BD Biosciences, San Jose, CA, USA) and isotype mouse IgG2_aκ (BD Biosciences, San Jose, CA, USA). For PD-L1 expression, cells were stained with anti-human CD274 or PD-L1 conjugated with PE-Cy7 monoclonal antibody clone MIHI (eBiosciences, Thermo Fisher Scientifics, Waltham, MA, USA)and isotype IgG₁κ (BD Biosciences, San Jose, CA, USA). For PD-1 expression, co-cultured cells were collected and stained with PE-conjugated anti-CD3 together with PE-Cy7-conjugated anti-PD1 antibodies (BD Biosciences, San Jose, CA, USA). For flow cytometric staining, cells were collected and washed with PBS containing 2% FBS, and then blocked with mouse and human serum at 4 °C for 30 min. Cells were incubated with antibodies according to the manufacturer's instructions. For each fluorochrome-labeled Ab used, appropriate isotype controls were included to establish a negative gate. After antibody staining, the cells were washed twice and fixed with 2% paraformaldehyde/PBS. Data acquisition and analysis were performed on an BD LSRII flow cytometer (BD Biosciences, San Jose CA, USA) or BD Accuri™ C6 Plus flow cytometer (BD Biosciences, San Jose CA, USA) using FACSDiva software (BD Biosciences, San Jose, CA, USA) or FlowJo software (version 7.1.3.0, Tree Star Inc., Padadena, TX, USA).

Sujjitjoon J., Sayour E., Tsao S.T., Uiprasertkul M., Sanpakit K., Buaboonnam J., Yenchitsomanus P.T., Atchaneeyasakul L.O, & Chang L.J. (2020). GD2-specific chimeric antigen receptor-modified T cells targeting retinoblastoma – assessing tumor and T cell interaction. Translational Oncology, 14(2), 100971.

+ Open protocol

+ Expand

Murine T Cell Immunophenotyping

Check if the same lab product or an alternative is used in the 5 most similar protocols

Peripheral blood of mice was collected on Day 45 after first immunization. Erythrocytes in collected blood were lysed with a 1× RBC lysis buffer (Invitrogen, Cat#00-4333-57). After washing with cold PBS, the cells were incubated with PE-conjugated anti-CD3, APC-conjugated anti-CD4- and PE-Cy7-conjugated anti-CD8 antibodies (BD, Franklin Lakes, NJ, USA) at 4 °C for 30 min. The cells were then resuspended in a PBS buffer containing 1% FBS and analyzed with a BD Celesta flow cytometer.

Luo Q., Ahmed W., Dai Y., Mohsin A., Hang H., Zhuang Y, & Guo M. (2021). Evaluation of a Virus-like Nanoparticle Porcine Circovirus Type-2 (PCV2) Capsid Protein Fused with the Pig Immunoglobulin Fc Fragment as a Novel Vaccine Candidate against PCV2 in Mice. Vaccines, 9(10), 1128.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!