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. 2019 Dec 19;9:436. doi: 10.3389/fcimb.2019.00436

Figure 3.

Figure 3

Western blots and confocal micrographs of chPD-1 and chPD-L1. (A) The extracellular soluble domains of chPD-1 and chPD-L1 were sub cloned into pkW06 expression vector and expressed as a human Fc-tagged protein, namely, pKW06-chPD-1-human-IgG1Fc and pKW06-chPD-L1-human-IgG1Fc. Proteins were resolved on an SDS 4–12% Bis-Tris gel along with 10% of input protein and western blotted with monoclonal anti-chPD-1 and chPD-L1 antibodies. Images demonstrate that ~55 kDa (red arrow left direction) for chPD-1 and a band of ~70 kDa (black arrow right direction) for chPD-L1. I is purified sample, II is supernatant from transfected cells and P is commercial positive control. Dividing black line indicates that intervening lanes have been spliced out. (B,C) Representative confocal micrographs illustrating chPD-1 and chPD-L1 in COS (left) and DF-1 (right) cells. Immunofluorescence staining was carried out on COS and DF-1 cells transfected with expression constructs containing pKW06-chPD-1-human-IgG1Fc and pKW06-chPD-L1-human-IgG1Fc plasmids. Cells were fixed after 24–48 h and stained with monoclonal anti-chPD-1 (A) and chPD-L1 (B) antibodies. Scale bar represents 40 μm.