Figure 4
Binding of affinity-purified human IgG molecules to nucleosomes. (a) Binding of purified DNase-I-treated antibodies to nucleosomes in a direct ELISA. The standard deviations were as follows: <0.26 OD for all points on the curve B3VH/B33VL, <0.45 OD for all points on the curve B3VH/B3VL, <0.33 OD for all points on the curve B3VH/B3(R27aS)VL, and <0.01 OD for all points on curve B3VH/BUVL. ODs in the negative control wells were all less than 0.07. (b)Binding to dsDNA of purified DNase-I-treated antibodies diluted in sample-enzyme-conjugate buffer containing nucleosomes at a concentration of 10 ÎĽg dsDNA/ml. At this concentration of nucleosomes, only B3VH/B3(R27aS)VL binds to dsDNA. The same results were obtained when the experiment was repeated at a nucleosome concentration of 2.5 ÎĽg dsDNA/ml, except that the peak OD for the B3VH/B3(R27aS)VL curve was lower. At nucleosome concentrations of 1.5 ÎĽg or 20 ÎĽg dsDNA/ml, none of these heavy/light combinations bound dsDNA. The standard deviation was <0.18 OD for all points on the curve B3VH/B3(R27aS)VL and <0.01 for all the points on all the other curves. ODs in the negative control wells were all less than 0.068.