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. 2017 Apr 19;8(31):51238–51252. doi: 10.18632/oncotarget.17228

Figure 3. Binding affinity and binding capacity of humanized antibodies.

Figure 3

(A) Affinity of cG7, hG7-BM1 and hG7-BM3 to CD24-GST analyzed by SPR. ka and kd of hG7-BM3 (Aiii) were 7.99×105 1/Ms and 4.55×10−4 1/s, and KD was 5.70×10−10 M. cG7 (Ai): ka was 1.76×1061/Ms, kd was 3.36×10−41/s, KD was 1.91×10−10 M. hG7-BM1 (Aii): ka was 4.65×1051/Ms, kd was 7.64×10−41/s, KD was 1.64×10−10 M. (B to F) Binding capacity of humanized antibodies to hepatoma cell lines. (Bi and Ci) cG7 exhibited considerable affinity with two tumor cells (Huh-7 for 88.4%, BEL-7402 for 79.3%). (Bii and Cii) Compared with cG7, hG7-BM1 showed lower binding rate with Huh-7 (58.2%) and BEL-7402 (54.5%). (Biii and Ciii) hG7-BM3 showed similar binding capacity to cG7 (Huh-7 for 77.9%, BEL-7402 for 69.1%). (Biv and Civ) Two hepatoma cell lines showed high expression levels of CD24 (94.1% in Huh-7 and 92.2% in BEL-7402). (Di to iv) These antibodies showed no binding affinity with normal human hepatic cell line HL-7702. (Ei, Eii and Fi, Fii) When the CD24 knockdown, the binding rates of hG7-BM3 to Huh-7 and BEL-7402 were reduced to 31.4% and 30.5%. (Eiii, Eiv and Fiii, Fiv) The efficiency of CD24 knockdown in Huh-7 and BEL-7402 was 55.3% and 55.7%.