human bcma fc chimera protein  (R&D Systems)

Journal: Antibodies

Article Title: Development of a Bispecific IgG1 Antibody Targeting BCMA and PDL1

doi: 10.3390/antib13010015

Figure Lengend Snippet: Structure and specificity of binding of chimeric BCMA×PDL1 bsAb. ( A ) Starting from the N-terminus, the fused heavy chain is composed of the anti-BCMA J22.9 antibody VH sequence-CH1 hinge1-A1linker-anti-PDL1 atezolizumab VH sequence-CH1 hinge2-CH2-CH3. The CH1 hinge-CH2 and CH3 sequences are from human IgG1. The two light chains (both k) are anti-BCMA J22.9 VL-CL and anti-PDL1 atezolizumab VL-CL. The red dot indicates the paired complementary mutations on CH1 and CL of the anti-PDL1 moiety to drive correct light chain pairing . ( B ) Specificity of binding of the purified bsAb and respective mAbs was tested by flow cytometry on mBCMA + and PDL1 + single-positive cell lines, KMS11 and HDLM2, respectively. MFI: mean fluorescence intensity. %: percentage of mBCMA-positive cells.

Article Snippet: After chip rotation, the following analytes, diluted in SPR running buffer (Dulbecco’s Phosphate-Buffered Saline with 0.005% Tween-20), were injected simultaneously on all the immobilized antibodies: recombinant human BCMA extracellular domain Fc chimera (recBCMA, R&D system, Minneapolis, MN, USA) and recombinant human PDL1 extracellular domain-Fc chimera (recPDL1, R&D system).

Techniques: Binding Assay, Sequencing, Purification, Flow Cytometry, Fluorescence

Journal: Antibodies

Article Title: Development of a Bispecific IgG1 Antibody Targeting BCMA and PDL1

doi: 10.3390/antib13010015

Figure Lengend Snippet: Relative binding affinity of BCMA×PDL1 bsAb and respective mAbs for target antigens. The relative affinity of the BCMA×PDL1 bsAb, anti-BCMA, and anti-PDL1 mAbs was tested by flow cytometry, using increasing concentrations of primary antibodies and detection with anti-human Fc-FITC secondary antibody. ( A ) Binding of bsAb and mAbs to mBCMA + KMS11 cell line. ( B ) Binding of bsAb and mAbs to PDL1 + HDM2 cell line. ( C ) Relative binding affinities (IC 50 ) for each antigen; NA: not applicable.

Article Snippet: After chip rotation, the following analytes, diluted in SPR running buffer (Dulbecco’s Phosphate-Buffered Saline with 0.005% Tween-20), were injected simultaneously on all the immobilized antibodies: recombinant human BCMA extracellular domain Fc chimera (recBCMA, R&D system, Minneapolis, MN, USA) and recombinant human PDL1 extracellular domain-Fc chimera (recPDL1, R&D system).

Techniques: Binding Assay, Flow Cytometry

Journal: Antibodies

Article Title: Development of a Bispecific IgG1 Antibody Targeting BCMA and PDL1

doi: 10.3390/antib13010015

Figure Lengend Snippet: Binding constants determined by SPR studies.

Article Snippet: After chip rotation, the following analytes, diluted in SPR running buffer (Dulbecco’s Phosphate-Buffered Saline with 0.005% Tween-20), were injected simultaneously on all the immobilized antibodies: recombinant human BCMA extracellular domain Fc chimera (recBCMA, R&D system, Minneapolis, MN, USA) and recombinant human PDL1 extracellular domain-Fc chimera (recPDL1, R&D system).

Techniques: Binding Assay

Journal: Antibodies

Article Title: Development of a Bispecific IgG1 Antibody Targeting BCMA and PDL1

doi: 10.3390/antib13010015

Figure Lengend Snippet: Surface plasmon resonance analysis. The sensorgrams shown were obtained by injecting recPDL1 or recBCMA, alone ( A – C ) or in succession ( D ) over immobilized anti-PDL1 ( A ), anti-BCMA ( B ), or BCMA×PDL1 bsAb ( C , D ). The antigens were flowed for 3 min, as indicated by the dashed lines.

Article Snippet: After chip rotation, the following analytes, diluted in SPR running buffer (Dulbecco’s Phosphate-Buffered Saline with 0.005% Tween-20), were injected simultaneously on all the immobilized antibodies: recombinant human BCMA extracellular domain Fc chimera (recBCMA, R&D system, Minneapolis, MN, USA) and recombinant human PDL1 extracellular domain-Fc chimera (recPDL1, R&D system).

Techniques: SPR Assay

Journal: Antibodies

Article Title: Development of a Bispecific IgG1 Antibody Targeting BCMA and PDL1

doi: 10.3390/antib13010015

Figure Lengend Snippet: The BCMA×PDL1 bsAb blocks APRIL binding to mBCMA and PD1-PDL1 interaction. ( A ) To test the ability of bsAb to block APRIL binding to mBCMA, we used CEM-mBCMA + cell line, increasing concentrations of bsAb and anti-BCMA mAb, a Flag-tagged APRIL protein, and an anti-Flag antibody. *: p < 0.05. ( B ) For assessing the inhibition of the PD1-PDL1 axis, we tested increasing concentrations of BCMA×PDL1 bsAb or anti-PDL1 mAb in the cell-based PD1/PDL1 Blockade Bioassay. ( C ) Cell-based PD1/PDL1 Blockade Bioassay in presence of recBCMA. Atezolizumab and cetuximab were used as positive and negative controls, respectively. *: p < 0.05 vs. PDL1.

Article Snippet: After chip rotation, the following analytes, diluted in SPR running buffer (Dulbecco’s Phosphate-Buffered Saline with 0.005% Tween-20), were injected simultaneously on all the immobilized antibodies: recombinant human BCMA extracellular domain Fc chimera (recBCMA, R&D system, Minneapolis, MN, USA) and recombinant human PDL1 extracellular domain-Fc chimera (recPDL1, R&D system).

Techniques: Binding Assay, Blocking Assay, Inhibition, Bioassay

Journal: Antibodies

Article Title: Development of a Bispecific IgG1 Antibody Targeting BCMA and PDL1

doi: 10.3390/antib13010015

Figure Lengend Snippet: The BCMA×PDL1 bsAb mediates CDC of mBCMA + cells. ( A ) The BJAB-mBCMA + cell line was incubated with increasing concentrations of bsAb, mAbs, or RTX as positive control and in the presence of 50% HS as a source of complement. CDC was measured after 4 h by 7-AAD staining and flow cytometry. *: p < 0.05 and **: p < 0.01. ( B ) Flow cytometry histograms showing the expression of mBCMA of BJAB cells stably expressing BCMA in presence or absence of DAPT. MFI: mean fluorescence intensity. %: percentage of mBCMA-positive cells.

Article Snippet: After chip rotation, the following analytes, diluted in SPR running buffer (Dulbecco’s Phosphate-Buffered Saline with 0.005% Tween-20), were injected simultaneously on all the immobilized antibodies: recombinant human BCMA extracellular domain Fc chimera (recBCMA, R&D system, Minneapolis, MN, USA) and recombinant human PDL1 extracellular domain-Fc chimera (recPDL1, R&D system).

Techniques: Incubation, Positive Control, Staining, Flow Cytometry, Expressing, Stable Transfection, Fluorescence