Fig. 1: Isolation of GPC1-specific antibodies using hybridoma technology and phage display technology. | Nature Communications

Fig. 1: Isolation of GPC1-specific antibodies using hybridoma technology and phage display technology.

From: The IgG4 hinge with CD28 transmembrane domain improves VHH-based CAR T cells targeting a membrane-distal epitope of GPC1 in pancreatic cancer

Fig. 1

a Discovery of anti-GPC1 monoclonal antibody (mAb) HM2 and camel VHH nanobody D4. The illustration was created with BioRender.com. b Six mouse mAbs (HM1 to HM6) at 1 μg/ml from three parental clones only bound to human GPC1, but not other human glypican members by ELISA. n = 1 independent experiment. c Monoclonal phage ELISA analysis of reactivity of D4 to human/mouse GPC1 as well as other human glypican members. n = 3 independent experiments. d Octet kinetic analysis for the interaction between HM2 or D4 and human GPC1. n = 1 independent experiment. e Cell surface GPC1 expression in GPC1-negative A431 cells, GPC1-positive pancreatic cancer cells T3M4, GPC1-knockout (KO) T3M4 cells, as well as GPC1-overexpressing cells 2B9. Data are representative of two independent experiments. White peaks represent the cell surface staining with isotype control. Red colored peaks and shaded gray peaks represent the cell surface staining of GPC1 using HM2 and D4, respectively, at 10 μg/ml. f Enlarged views of a 2D class average of GPC1 in complex with HM2 Fab and GPC1 in complex with D4-LR. g A ribbon diagram of the GPC1/HM2 Fab/D4 model. D4 and HM2 recognize the membrane-distal and membrane-proximal epitopes of GPC1, respectively, which are on the opposite sides of GPC1. The epitopes are highlighted in red color. Data are represented as mean ± SEM. Source data are provided as a Source Data file.

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