Murine Anti-PD-L1 mAb

InvivoGen’s engineered Anti-PD-L1-mIgG1e3 InvivoFit™ antibody

Recombinant mAb against murine PD-L1 for in vivo use

Anti-PD-L1-mIgG1e3 InvivoFit™ is a recombinant monoclonal antibody (mAb) designed for in vivo studies in mice. It features the variable region of the anti-PD-L1 atezolizumab [1] and the engineered murine IgG1e3 constant region.  Programmed cell death ligand 1 (PD-L1; also called B7-H1 or CD274) is a transmembrane protein expressed on antigen-presenting cells. PD-L1 binds to programmed cell death protein 1 (PD-1) on T cells and contributes to T cell exhaustion during chronic infections. Moreover, it has been shown that the engagement of PD-1 on T cells by PD-L1 on tumor cells is associated with the immune escape of tumors. Clinical trials have highlighted the anti-tumor efficacy of blockades targeting the PD-1/PD-L1 pathway [2].  

Atezolizumab (formerly known as MPDL3280A) is a therapeutic mAb that binds human and murine PD-L1, blocking the interaction with its receptor programmed cell death 1 (PD-1). Notably, atezolizumab contains an engineered constant region designed to limit effector functions, such as antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) [3]. However, as it is a humanized antibody, it is immunogenic in mice. To overcome this issue, Anti-PD-L1-mIgG1e3 InvivoFit™ was generated by recombinant DNA technology so that it is ~65% murine (constant region). Importantly, its constant region contains a point mutation D265A (a replacement of aspartic acid by alanine at position 265), resulting in the complete loss of cytolytic effector functions [4].

Anti-PD-L1-mIgG1e3 is provided in an InvivoFit™ grade, a high-quality standard specifically adapted to in vivo studies.

Key features of Anti-PD-L1-mIgG1e3 InvivoFit™:

• Derived from Atezolizumab (aka MPDL3280A), human IgG1
• Feature mouse IgG1e3 isotype (constant region)
• mIgG1e3 (IgG1 with a D265A point mutation) is effectorless
• Filter-sterilized (0.2 µm), endotoxin level: 1 EU/mg
• Suitable for parenteral delivery in mice (azide-free)
• Low aggregation < 5%
• Produced in animal-free facilities and defined media

Anti-PD-L1-mIgG1e3 InvivoFit™ is produced in Chinese hamster ovary (CHO) cells and purified by affinity chromatography with protein A.  Its binding is validated by flow cytometry and ELISA.

References:

1. Lesniak W.G. et al., 2016. PD-L1 Detection in Tumors Using [(64)Cu]Atezolizumab with PET. Bioconjug Chem. 27(9):2103-10.
2. McDermott D. & Atkins M. 2013. PD-1 as a potential target in cancer therapy. Cancer Med. 2(5): 662–673.
3. Herbst R.S. et al., 2014. Predictive correlates of response to the anti-PD-L1 antibody MPDL3280A in cancer patients. Nature. 515(7528):563-7.
4. Baudino L. et al., 2008. Crucial role of aspartic acid at position 265 in the CH2 domain for murine IgG2a and IgG2b Fc-associated effector functions. J Immunol. 181(9):6664-9.

Figures

ELISA binding of Anti-PD-L1-mIgG1e3 InvivoFit™ to recombinant PD-L1. 

A dilution series of Anti-PD-L1-mIgG1e3 InvivoFit™ and anti-PD-L1 clone 10F-9G2 was used for the detection of coated murine PD-L1 (A) or human PD-L1 (B) antigen at 2 µg/ml. Bound antibody was detected using either an HRP-conjugated anti-murine IgG secondary antibody for Anti-PD-L1-mIgG1 e3 InvivoFit™ or an HRP-conjugated anti-rat IgG2b secondary antibody for anti-PD-L1 clone 10F-9G2. HRP activity was assessed by a colorimetric assay using Tetramethylbenzidine (TMB) substrate. Optical density was measured at 490 nm. Data are shown as percentage absorbance.

Specifications

Specificity: Targets cells expressing murine or human PD-L1

Formulation: Lyophilized from 0.2 μm filtered solution in 150 mM sodium chloride, 20 mM sodium phosphate buffer with 5% saccharose

Clonality: Monoclonal antibody

Isotype: Murine IgG1e3 (D265A mutation; no effector function), kappa

Control: mIgG1e3 InvivoFit™ isotype control

Source: CHO cells

Purity: Purified by affinity chromatography with protein A

Tested applications: Flow cytometry and ELISA

Quality control:

• Binding confirmed by flow cytometry
• The complete sequence of this antibody has been verified
• < 5% aggregates (confirmed by size exclusion chromatography)
• Endotoxin level <1 EU/mg (determined by the LAL assay)

Contents

Anti-PD-L1-mIgG1e3 InvivoFit™ is Filter-sterilized (0.2 µm), endotoxin-free, azide-free, and lyophilized.

This product is available in three pack sizes:

• pdl1-mab15-1 : 1 mg
• pdl1-mab15-10 : 10 mg
• pdl1-mab15-50 : 50 mg (5 x 10 mg)

 The product is shipped at room temperature.

 Store lyophilized antibody at -20 °C.

 Lyophilized product is stable for at least 1 year.

 Avoid repeated freeze-thaw cycles.

InvivoFit™

InvivoFit™ is a high-quality standard specifically adapted for in vivo studies. InvivoFit™ products are filter-sterilized (0.2 µm) and filled under strict aseptic conditions in a clean room. The level of bacterial contaminants (endotoxins and lipoproteins) in each lot is verified using a LAL assay and a TLR2 and TLR4 reporter assay.