Generation and Purification of PRLR-DbsAb

The PRLR-DbsAb was generated by BAPTS system, which was previously described in detail [27 (link),28 (link)]. CD3 antibody-fusion protein (Fragment A) was expressed in a stably transfected cell line CHO while PRLR antibody-fusion protein (Fragment B) was expressed in 293E cells using transient gene expression (TGE) technology. Both Fragment A and Fragment B were purified by Protein L affinity chromatography (GE Healthcare). The monoclonal antibodies used in this study including PRLR antibody (PRLR mAb), CD3 antibody (CD3 mAb) and PD-1 antibody (PD-1 mAb) were expressed by 293E cells and purified by Protein A affinity chromatography (GE Healthcare). The affinity of CD3 mAb and PD-1 mAb to their antigens was previously confirmed [27 (link),29 (link)]. The amino acid sequence used for the PRLR mAb is identical to that of the PRLR monoclonal antibody LFA102 [5 (link)]. All recombinant antibodies were dialyzed overnight to phosphate buffer saline (PBS) and sterilized by filtration using a 0.22 μm filter.

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Zhou Y., Zong H., Han L., Xie Y., Jiang H., Gilly J., Zhang B., Lu H., Chen J., Sun R., Pan Z, & Zhu J. (2020). A novel bispecific antibody targeting CD3 and prolactin receptor (PRLR) against PRLR-expression breast cancer. Journal of Experimental & Clinical Cancer Research : CR, 39, 87.

Publication 2020

Protein L affinity chromatography Protein A affinity chromatography

Affinity chromatography Amino acid sequence Antibodies Antibody Antibody affinity Antigens Buffer Cell line Cells Filtration Gene expression Lfa102 monoclonal antibody Monoclonal antibodies Phosphate Prlr Protein Protein a Saline Transient

Corresponding Organization :

Other organizations : Shanghai Jiao Tong University

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Variable analysis

independent variables

Expression system (CHO cells for Fragment A, 293E cells for Fragment B)
Purification method (Protein L affinity chromatography for Fragments A and B, Protein A affinity chromatography for PRLR mAb, CD3 mAb, and PD-1 mAb)

dependent variables

Not explicitly mentioned

control variables

Antibody types (PRLR mAb, CD3 mAb, PD-1 mAb)
Amino acid sequence of PRLR mAb (identical to LFA102)
Dialysis to PBS and sterilization by 0.22 μm filtration for all recombinant antibodies

positive controls

Affinity of CD3 mAb and PD-1 mAb to their antigens, previously confirmed [27, 29]

negative controls

Not explicitly mentioned

Annotations

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