polyclonal goat anti human bcma ab  (R&D Systems)

Journal: Assay and drug development technologies

Article Title: A Flexible Multiplatform Bioanalytical Strategy for Measurement of Total Circulating Shed Target Receptors: Application to Soluble B Cell Maturation Antigen Levels in the Presence of a Bispecific Antibody Drug.

doi: 10.1089/adt.2020.1024

Figure Lengend Snippet: Fig. 1. Mechanism of drug action and impact of sBCMA. (A) BCMA CD3 bispecific antibody binds to membrane-bound BCMA on MM cells and CD3 on T cells. (B) sBCMA in the central compartment can compete with membrane-bound BCMA for binding to the drug, thereby producing a sink that can reduce the efficacious drug concentration. Levels of the soluble form may also be used as a biomarker for disease progression. BCMA, B cell maturation antigen; MM, multiple myeloma; sBCMA, soluble BCMA.

Article Snippet: Polyclonal goat anti-human BCMA antibody (R&D Systems) and polyclonal rabbit anti-peptide (CSSTPPLTCQR) antibodies (Cambridge Research Biochemicals) were buffer exchanged into phosphate-buffered saline (PBS), pH 7.4 (Gibco, Grand Island, NY) using a Zeba spin desalting column, 40 k MWCO (Thermo Scientific) following the manufacturer’s instructions.

Techniques: Membrane, Binding Assay, Concentration Assay, Biomarker Discovery

Journal: Assay and drug development technologies

Article Title: A Flexible Multiplatform Bioanalytical Strategy for Measurement of Total Circulating Shed Target Receptors: Application to Soluble B Cell Maturation Antigen Levels in the Presence of a Bispecific Antibody Drug.

doi: 10.1089/adt.2020.1024

Figure Lengend Snippet: Fig. 2. BCMA CD3 bispecific antibody interferes extensively with measurement of total sBCMA by LBA. Four concentrations of BCMA CD3 bispecific antibody (blue squares =no spike, red triangle = 0.069 nM, inverted green triangle = 0.55 nM, purple diamond =6.9 nM, orange circle = 55 nM) were spiked into: (A) QCs prepared by spiking recombinant monkey sBCMA into surrogate matrix at 75.2, 8.84, 1.04, 0.12 nM (denoted HQC, MQC, LQC, and LLOQ, respectively) and (B) four naive cynomolgus monkey plasma samples with endogenous sBCMA. As the drug concentrations increased, the re- covered amount of sBCMA rapidly decreased. HQC, high quality control; LBA, ligand binding assay; LLOQ, lower limit of quantitation; LQC, low quality control; MQC, mid quality control.

Article Snippet: Polyclonal goat anti-human BCMA antibody (R&D Systems) and polyclonal rabbit anti-peptide (CSSTPPLTCQR) antibodies (Cambridge Research Biochemicals) were buffer exchanged into phosphate-buffered saline (PBS), pH 7.4 (Gibco, Grand Island, NY) using a Zeba spin desalting column, 40 k MWCO (Thermo Scientific) following the manufacturer’s instructions.

Techniques: Recombinant, Clinical Proteomics, Control, Ligand Binding Assay, Quantitation Assay

Journal: Assay and drug development technologies

Article Title: A Flexible Multiplatform Bioanalytical Strategy for Measurement of Total Circulating Shed Target Receptors: Application to Soluble B Cell Maturation Antigen Levels in the Presence of a Bispecific Antibody Drug.

doi: 10.1089/adt.2020.1024

Figure Lengend Snippet: Fig. 3. Protein-level IA-LC-MS/MS workflow and minimization of drug interference. (A) sBCMA was captured with biotinylated polyclonal anti-BCMA antibodies, which were immunoprecipitated using streptavidin-coated magnetic beads. Beads were washed, bound sBCMA was eluted, and eluates were processed to generate tryptic peptides that were analyzed by LC-MS/MS. (B) Suppression of peak area in the presence of BCMA CD3 bispecific antibody relative to the unspiked condition was overcome by titrating the capture antibody. IA, im- munoaffinity; LC-MS/MS, liquid chromatography–tandem mass spectrometry.

Article Snippet: Polyclonal goat anti-human BCMA antibody (R&D Systems) and polyclonal rabbit anti-peptide (CSSTPPLTCQR) antibodies (Cambridge Research Biochemicals) were buffer exchanged into phosphate-buffered saline (PBS), pH 7.4 (Gibco, Grand Island, NY) using a Zeba spin desalting column, 40 k MWCO (Thermo Scientific) following the manufacturer’s instructions.

Techniques: Liquid Chromatography with Mass Spectroscopy, Immunoprecipitation, Magnetic Beads, Liquid Chromatography, Mass Spectrometry

Journal: Assay and drug development technologies

Article Title: A Flexible Multiplatform Bioanalytical Strategy for Measurement of Total Circulating Shed Target Receptors: Application to Soluble B Cell Maturation Antigen Levels in the Presence of a Bispecific Antibody Drug.

doi: 10.1089/adt.2020.1024

Figure Lengend Snippet: Fig. 5. Total sBCMA levels in cynomolgus monkeys dosed with bispecific antibody and in un- treated human MM patients. (A) Concentration of total plasma sBCMA increases after dosing with BCMA CD3 bispecific antibody (blue circle = 1 mg/kg, red square = 10 mg/kg, and green triangle = 100 mg/kg). (B) Concentration of sBCMA in plasma samples from relapsed/refractory MM patients. sBCMA level in pooled normal plasma sample depicted by dotted line (1.75 nM).

Article Snippet: Polyclonal goat anti-human BCMA antibody (R&D Systems) and polyclonal rabbit anti-peptide (CSSTPPLTCQR) antibodies (Cambridge Research Biochemicals) were buffer exchanged into phosphate-buffered saline (PBS), pH 7.4 (Gibco, Grand Island, NY) using a Zeba spin desalting column, 40 k MWCO (Thermo Scientific) following the manufacturer’s instructions.

Techniques: Concentration Assay, Clinical Proteomics

Journal: Assay and drug development technologies

Article Title: A Flexible Multiplatform Bioanalytical Strategy for Measurement of Total Circulating Shed Target Receptors: Application to Soluble B Cell Maturation Antigen Levels in the Presence of a Bispecific Antibody Drug.

doi: 10.1089/adt.2020.1024

Figure Lengend Snippet: Fig. 3. Protein-level IA-LC-MS/MS workflow and minimization of drug interference. (A) sBCMA was captured with biotinylated polyclonal anti-BCMA antibodies, which were immunoprecipitated using streptavidin-coated magnetic beads. Beads were washed, bound sBCMA was eluted, and eluates were processed to generate tryptic peptides that were analyzed by LC-MS/MS. (B) Suppression of peak area in the presence of BCMA CD3 bispecific antibody relative to the unspiked condition was overcome by titrating the capture antibody. IA, im- munoaffinity; LC-MS/MS, liquid chromatography–tandem mass spectrometry.

Article Snippet: Biotinylated goat polyclonal anti-BCMA (DY193; R&D Systems) was added to each standard, QC, and samples followed by fourfold dilution in Super Block and then incubated at 4 C overnight on a rotator.

Techniques: Liquid Chromatography with Mass Spectroscopy, Immunoprecipitation, Magnetic Beads, Liquid Chromatography, Mass Spectrometry

Journal: Blood Advances

Article Title: Single- and double-hit events in genes encoding immune targets before and after T cell–engaging antibody therapy in MM

doi: 10.1182/bloodadvances.2021004418

Figure Lengend Snippet: WGS, read coverage track, and immunohistochemistry. (A) Circos plot with copy number variations, structural variations, and single-nucleotide variants based on WGS data at progression after therapy with CD3xBCMA BsAb. Outer track runs clockwise from chromosome 1 to Y. Inner track shows gains >1 Mb in blue and losses >1 Mb in red. Red lines inside the circle represent interchromosomal translocations with variant allele frequency (VAF) >0.1. Genes with mutations (CRBN) are depicted in red. (B) Read coverage visualized with Integrative Genomics Viewer illustrates copy number variations in the short arm of chromosome 16 comprising heterozygous deleted regions caused by monosomy 16 and a homozygous deletion of 586 kb from 12.036.001 to 12.622.000 (hg19 reference genome) including TNFRSF17 (BCMA ). (C) BCMA protein expression was determined by immunohistochemistry using a polyclonal goat anti-BCMA antibody on paraffin-embedded bone marrow sections. Lambda light chain staining (immunohistochemistry) served as positive control (magnification, x200).

Article Snippet: Immunohistochemistry BCMA protein expression was determined by immunohistochemistry using a polyclonal goat anti-BCMA antibody (dilution 1:10; target retrieval pH 6.1) (AF193, R&D Systems) on paraffin-embedded bone marrow sections according to standard procedures.

Techniques: Immunohistochemistry, Variant Assay, Expressing, Staining, Positive Control