Human Anti-MS4A1 Recombinant Antibody (clone OFA) (CAT#: HPAB-N0181-YC)

Provided is a monoclonal antibody that has specificity affinity for CD20. It can be a special property combination of CD20 antigen molecules, so as to single-minded kill B lymphoma cells.


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Figure 1 OFA-opsonized B cells bind more C1q than RTX-opsonized B cells.

Figure 1 OFA-opsonized B cells bind more C1q than RTX-opsonized B cells.

A and B, Varying amounts of C1q were added to B cells suspended in complete RPMI 1640 medium and then reacted with either Al647 OFA, Al647 RTX, or Al647 7D8-IgG4 (K322A). C, Binding of C1q to CLL cells opsonized with either OFA or with RTX, similar conditions and analyses. D-I, Cells were combined in cold medium with Al647 mAb and varying amounts of C1q and then incubated at 37°C. Aliquots were removed at the indicated times, quenched with ice-cold BSA-PBS, washed, probed with FITC anti-C1q, and then analyzed by flow cytometry for C1q binding (D-F) and mAb binding (G-I). D and G, Raji cells. E and H, Daudi cells. F and I, Z138 cells.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

IF

Figure 2 C1q colocalizes with bound OFA on Daudi cells.

Figure 2 C1q colocalizes with bound OFA on Daudi cells.

A, Fluorescence intensity of Al488 C1q and Al647 mAbs bound to Daudi cells. Very little C1q binding to RTX- or 7D8-IgG4 (K322A)-opsonized cells was observed. B and C, Fluorescence signals for Al647 OFA-opsonized samples were analyzed for colocalization with bound Al488 C1q.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

Figure 3 OFA-opsonized cells are killed more effectively in C1q depleted serum supplemented with low C1q concentrations than are RTX opsonized cells.

Figure 3 OFA-opsonized cells are killed more effectively in C1q depleted serum supplemented with low C1q concentrations than are RTX opsonized cells.

A, Daudi cells were combined with OFA, RTX, or with no mAb in 50% C1q-depleted NHS supplemented with varying amounts of C1q. B and C, In the presence of C1q, OFA-opsonized cells are killed very rapidly. OFA-opsonized Raji (B) or Z138 (C)cells were incubated at 37°C for 1-20 min in 50% C1q-depleted NHS supplemented with 1, 2, or 35 μg/ml C1q. D and E, In C1q-depleted NHS supplemented with either 35 or 1 μg/ml C1q, binding of C1q to OFA opsonized Raji or Z138 cells is quite weak.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

Figure 4 In C1q-depleted serum supplemented with limited amounts of C1q, OFA-opsonized Raji cells have higher levels of C3b deposition and undergo greater CDC than do RTX-opsonized Raji cells.

Figure 4 In C1q-depleted serum supplemented with limited amounts of C1q, OFA-opsonized Raji cells have higher levels of C3b deposition and undergo greater CDC than do RTX-opsonized Raji cells.

A, In medium, OFA-opsonized Raji cells bind more C1q than do RTX-opsonized cells. B and C, C3b deposition and CDC as a function of time and C1q concentration.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

Figure 5 In C1q-depleted serum supplemented with limited amounts of C1q, OFA-opsonized Daudi cells have higher levels of C3b deposition and undergo greater CDC than do RTX-opsonized Daudi cells.

Figure 5 In C1q-depleted serum supplemented with limited amounts of C1q, OFA-opsonized Daudi cells have higher levels of C3b deposition and undergo greater CDC than do RTX-opsonized Daudi cells.

A, In medium, OFA-opsonized Daudi cells bind more C1q than do RTX-opsonized cells. B and C, C3b deposition and CDC as a function of time and C1q concentration.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

Figure 6 In C1q-depleted serum supplemented with limited amounts of C1q, OFA-opsonized Z138 cells have higher levels of C3b deposition and undergo greater CDC than do RTX-opsonized Z138 cells.

Figure 6 In C1q-depleted serum supplemented with limited amounts of C1q, OFA-opsonized Z138 cells have higher levels of C3b deposition and undergo greater CDC than do RTX-opsonized Z138 cells.

A, In medium, OFA-opsonized Z138 cells bind more C1q than do RTX-opsonized cells. B and C, C3b deposition and CDC as a function of time and C1q concentration.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

Figure 7 In C1q-depleted serum supplemented with limited amounts of C1q, primary CLL cells opsonized with OFA bind more C1q than cells opsonized with RTX, have higher levels of C3b deposition, and have more CDC.

Figure 7 In C1q-depleted serum supplemented with limited amounts of C1q, primary CLL cells opsonized with OFA bind more C1q than cells opsonized with RTX, have higher levels of C3b deposition, and have more CDC.

A-C, Dose-response profile for CLL cells from one CLL patient. D, Results for C1q binding in medium, and C3b deposition and CDC in C1qdepleted serum (35 μg/ml C1q for all cases) for CLL cells from seven patients.

Pawluczkowycz, A. W., Beurskens, F. J., Beum, P. V., Lindorfer, M. A., van de Winkel, J. G., Parren, P. W., & Taylor, R. P. (2009). Binding of submaximal C1q promotes complement-dependent cytotoxicity (CDC) of B cells opsonized with anti-CD20 mAbs ofatumumab (OFA) or rituximab (RTX): considerably higher levels of CDC are induced by OFA than by RTX. The Journal of Immunology, 183(1), 749-758.

Figure 8 OFA induces greater cell kill through CDC compared with RTX.

Figure 8 OFA induces greater cell kill through CDC compared with RTX.

OFA demonstrated much greater cell kill by CDC than by RTX, with 20 mg/ml eliciting high levels of cell death in all cytogenetic groups. Statistical analysis revealed that, when compared with RTX, OFA induced significantly more CDCmediated cell death in 11q - and 17p- cohorts, with borderline significant effect in the normal/13q group

Middleton, O., Cosimo, E., Dobbin, E., McCaig, A. M., Clarke, C., Brant, A. M., ... & Wheadon, H. (2015). Complement deficiencies limit CD20 monoclonal antibody treatment efficacy in CLL. Leukemia, 29(1), 107-114.

Figure 9 Binding stability of CD20 mAbs to human lymphoma B cell lines.

Figure 9 Binding stability of CD20 mAbs to human lymphoma B cell lines.

Binding of 60 nM Ofatumumab (OFA), Rituximab (RTX), or Obinutuzumab (OBI) to Daudi (A), Ramos (B), LCL1.11 (C), or P493.6 (D) cells was recorded until equilibrium was approached (not shown) followed by measurement of mAb dissociation either in plain cell culture medium or in presence of 60 nM of the respective unlabeled (unlab.) antibody. For all cell lines the dissociation of OFA was measured in presence of 180 nM unlabeled OFA (instead of 60 nM) to enhance possible cell-line differences. (E) Signal intensities were normalized to 100% at the beginning of the dissociation. The remaining signal after 1 h (black) and 2 h (gray) dissociation both in plain media (solid) and in presence of unlabeled antibody (shaded) are plotted for human B cell lines.

Bondza, S., Marosan, A., Kara, S., Lösing, J., Peipp, M., Nimmerjahn, F., ... & Lux, A. (2021). Complement-dependent activity of CD20-specific IgG correlates with bivalent antigen binding and C1q binding strength. Frontiers in immunology, 11, 609941.

Figure 10 Binding stability of CD20 mAbs to human primary B cells.

Figure 10 Binding stability of CD20 mAbs to human primary B cells.

Binding of 60 nM Ofatumumab (OFA), Rituximab (RTX) or Obinutuzumab (OBI) to primary human B cells isolated from blood of healthy donors was recorded until equilibrium was approached (not shown) and then mAb dissociation was measured either in plain cell culture medium or in presence of 60 nM of the respective unlabeled antibody. (A) One out of three independent measurements shown. (B) Signal intensities were normalized to 100% at the beginning of the dissociation. The remaining signal after 1 h (black) and 2 h (gray) dissociation both in plain media (solid) and in presence of unlabeled antibody (shaded) are plotted.

Bondza, S., Marosan, A., Kara, S., Lösing, J., Peipp, M., Nimmerjahn, F., ... & Lux, A. (2021). Complement-dependent activity of CD20-specific IgG correlates with bivalent antigen binding and C1q binding strength. Frontiers in immunology, 11, 609941.

Figure 11 Correlative studies on patients for the first 30 days of the trial.

Figure 11 Correlative studies on patients for the first 30 days of the trial.

OFA was infused on days 1, 8 and 29. Blood samples were obtained immediately before, and 2, 6 and 24 hrs after starting OFA infusions. Results are normalized to pre-treatment values for absolute lymphocyte counts (ALC), CD20, and CH50. Bound OFA is normalized to the 2 hr mark (first infusion), usually the maximum amount bound; values for bound C3d are normalized to maximum amount bound, observed at 2 or 6 hrs. Absolute values for these parameters are provided in Table I, along with representative uncertainties (SD). A. ALC; B. B cell surface levels of CD20; C. cell-bound OFA; D. complement titers (CH50 determinations); E. C3d deposition on B cells. The complement titer of patient N1 was low throughout the study and is not plotted in panel D. Results in B, C and E are based on duplicate determinations; CH50 titers were determined in triplicate.

Beurskens, F. J., Lindorfer, M. A., Farooqui, M., Beum, P. V., Engelberts, P., Mackus, W. J., ... & Taylor, R. P. (2012). Exhaustion of cytotoxic effector systems may limit monoclonal antibody-based immunotherapy in cancer patients. The Journal of Immunology, 188(7), 3532-3541.

Figure 12 The Serum Ofatumumab (OFA) Level at 8 Hours After Starting Therapy is Inversely Proportional to the Total Pre-treatment Cell Associated Circulating CD20 and the Decrease in this Value after the Initial Dose of 300 mg of OFA.

Figure 12 The Serum Ofatumumab (OFA) Level at 8 Hours After Starting Therapy is Inversely Proportional to the Total Pre-treatment Cell Associated Circulating CD20 and the Decrease in this Value after the Initial Dose of 300 mg of OFA.

These correlations showed that 8hr OFA levels were inversely correlated with pre-treatment total cell-associated CD20 and with the change in the total cellassociated CD20 between the 0 hr and 8 hr specimens.

Baig, N. A., Taylor, R. P., Lindorfer, M. A., Church, A. K., LaPlant, B. R., Pettinger, A. M., ... & Zent, C. S. (2014). Induced resistance to ofatumumab-mediated cell clearance mechanisms, including complement-dependent cytotoxicity, in chronic lymphocytic leukemia. The Journal of Immunology, 192(4), 1620-1629.

Figure 13 Levels of Ofatumumab (OFA) Binding to CLL Cells Equivalent to Those Measured in Circulating CLL Cells After Initiation of OFA Therapy Have Low in Vitro Complement Activating Activity.

Figure 13 Levels of Ofatumumab (OFA) Binding to CLL Cells Equivalent to Those Measured in Circulating CLL Cells After Initiation of OFA Therapy Have Low in Vitro Complement Activating Activity.

There was a progressive increase in OFA binding as the OFA concentration was increased from 0.025 μg/ml to 0.100 μg/ml.

Baig, N. A., Taylor, R. P., Lindorfer, M. A., Church, A. K., LaPlant, B. R., Pettinger, A. M., ... & Zent, C. S. (2014). Induced resistance to ofatumumab-mediated cell clearance mechanisms, including complement-dependent cytotoxicity, in chronic lymphocytic leukemia. The Journal of Immunology, 192(4), 1620-1629.


Specifications

  • Host Species
  • Human
  • Type
  • Human IgG
  • Specificity
  • Human MS4A1
  • Species Reactivity
  • Human
  • Clone
  • OFA
  • Applications
  • FC, FuncS

Product Property

  • Purity
  • >95% as determined by SDS-PAGE and HPLC analysis
  • Buffer
  • PBS
  • Preservative
  • No preservatives
  • Storage
  • Centrifuge briefly prior to opening vial. Store at +4°C short term (1-2 weeks). Aliquot and store at -20°C long term. Avoid repeated freeze/thaw cycles.

Target

  • Alternative Names
  • Membrane Spanning 4-Domains A1; Membrane-Spanning 4-Domains, Subfamily A, Member 1; Leukocyte Surface Antigen Leu-16; CD20 Antigen; CD20; Bp35; Membrane-Spanning 4-Domains Subfamily A Member 1; B-Lymphocyte Cell-Surface Antigen B1; B-Lymphocyte Surface Antigen B1

Product Notes

This is a product of Creative Biolabs' Hi-Affi™ recombinant antibody portfolio, which has several benefits including:

• Increased sensitivity
• Confirmed specificity
• High repeatability
• Excellent batch-to-batch consistency
• Sustainable supply
• Animal-free production

See more details about Hi-Affi™ recombinant antibody benefits.

Downloads

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See other products for "Clone OFA"

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Humanized Antibody

Hyper-galactosylated

High-mannose Glycoform

CAT Product Name Application Type
Gly-155LC-1 Recombinant Anti-Human MS4A1 Antibody (Fc glycosylation/High-mannose glycosylated) ELISA Humanized antibody

Deglycosylated Antibody (Non-glycosylated IgGs)

CAT Product Name Application Type
Gly-171LC Recombinant Anti-Human MS4A1 Antibody (Non-glycosylated) ELISA Chimeric antibody (mouse/human)

Neutralizing Antibody

CAT Product Name Application Type
NEUT-1751CQ Mouse Anti-MS4A1 Recombinant Antibody (clone CBLS-211) Neut, FuncS Mouse IgG1, κ

Rabbit Monoclonal Antibody

ADCC Enhanced Antibody

CAT Product Name Application Type
AFC-TAB-772 Afuco™ Anti-MS4A1 ADCC Recombinant Antibody (Ocrelizumab), ADCC Enhanced ELISA, IP, FC, FuncS, Neut, IF ADCC enhanced antibody
AFC-TAB-016 Afuco™ Anti-MS4A1 ADCC Recombinant Antibody (Rituximab), ADCC Enhanced IF, IP, Neut, FuncS, ELISA, FC ADCC enhanced antibody
AFC-TAB-028 Afuco™ Anti-MS4A1 ADCC Recombinant Antibody (Ofatumumab), ADCC Enhanced FC, IP, ELISA, Neut, FuncS, IF ADCC enhanced antibody
AFC-TAB-207 Afuco™ Anti-MS4A1 ADCC Recombinant Antibody (Ublituximab), ADCC Enhanced ELISA, IP, FC, FuncS, Neut, IF ADCC enhanced antibody
AFC-TAB-771 Afuco™ Anti-MS4A1 ADCC Recombinant Antibody (Ocaratuzumab), ADCC Enhanced IF, IP, Neut, FuncS, ELISA, FC ADCC enhanced antibody

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