immunohistochemistry mab anti rat cd2  (Bio-Rad)

Journal: Frontiers in Immunology

Article Title: Siplizumab, an Anti-CD2 Monoclonal Antibody, Induces a Unique Set of Immune Modulatory Effects Compared to Alemtuzumab and Rabbit Anti-Thymocyte Globulin In Vitro

doi: 10.3389/fimmu.2020.592553

Figure Lengend Snippet: Fc γ receptor (FcγR) signaling and complement-dependent cytolysis (CDC) assays Jurkat reporter cells (Promega) stably expressing Fc γ receptor (FcγR) I (A) , FcγRIIA (B) , or FcγRIIIA (C) , respectively, incubated with increasing concentrations of siplizumab or Fc-silent siplizumab. Reporter cells binding the Fc-fragment of a target-bound IgG antibody with their FcγR induced expression of a luciferase reporter gene resulting in a luminescence signal upon addition of assay substrate (Promega). Luminescence was normalized to highest concentration of siplizumab and is displayed as mean percentage ± SD (N = 3). (D) CDC of PBMC mixed with different doses of siplizumab or positive anti-CD2 control (BTI-322). Data displayed as mean percentage ± SD (N = 2).

Article Snippet: PBMC were isolated via density gradient centrifugation and incubated with siplizumab or positive control rat anti-CD2 IgG2b mAb (BTI-322, BioTransplant Inc, MA, USA) in 10% ultra-low IgG FBS in PBS at the indicated concentrations for 30 min at 1 × 10 7 cells per ml at RT with shaking.

Techniques: Stable Transfection, Expressing, Incubation, Binding Assay, Luciferase, Concentration Assay, Control

Journal: Frontiers in Immunology

Article Title: Siplizumab, an Anti-CD2 Monoclonal Antibody, Induces a Unique Set of Immune Modulatory Effects Compared to Alemtuzumab and Rabbit Anti-Thymocyte Globulin In Vitro

doi: 10.3389/fimmu.2020.592553

Figure Lengend Snippet: Target antigen expression on PBMC. Data is displayed as the average median fluorescent intensity (MFI) ± SD (N = 5). PBMC were stained using the following drugs: siplizumab (Target antigen CD2), Alemtuzumab (CD52) and rATG (Thymocyte antigens). After removal of unbound antibody, samples were stained with a secondary anti-human IgG antibody and MFI for the secondary antibody was recorded, representing the expression of the respective transplant antigens. Tcm, central memory T cells; Tem, effector memory T cells; Temra, effector-memory CD45RA + T cells; rTreg, resting regulatory T cells; nsTreg, non-suppressive Treg; aTreg, activated Treg; CD56 Bright , CD56 bright NK cells; CD56 Dim , CD56 dim NK cells; CD56 neg CD16 + , CD56 − CD16 + NK cells.

Article Snippet: PBMC were isolated via density gradient centrifugation and incubated with siplizumab or positive control rat anti-CD2 IgG2b mAb (BTI-322, BioTransplant Inc, MA, USA) in 10% ultra-low IgG FBS in PBS at the indicated concentrations for 30 min at 1 × 10 7 cells per ml at RT with shaking.

Techniques: Expressing, Staining

Journal: Frontiers in Immunology

Article Title: Siplizumab, an Anti-CD2 Monoclonal Antibody, Induces a Unique Set of Immune Modulatory Effects Compared to Alemtuzumab and Rabbit Anti-Thymocyte Globulin In Vitro

doi: 10.3389/fimmu.2020.592553

Figure Lengend Snippet: Summary of the characteristics of different tested compounds.

Article Snippet: PBMC were isolated via density gradient centrifugation and incubated with siplizumab or positive control rat anti-CD2 IgG2b mAb (BTI-322, BioTransplant Inc, MA, USA) in 10% ultra-low IgG FBS in PBS at the indicated concentrations for 30 min at 1 × 10 7 cells per ml at RT with shaking.

Techniques: Activation Assay