Not all these regions exhibited equal agglutinating activity, as evidenced by titer and the extent of the active areas. In all cases, most of the activity lay in the region of high anionic binding capacity. This was particularly noticeable in group A and group B sera, in which cases activity in Regions 1 and 2 was usually not detectable without prior concentration and occasionally could not be detected at all. There appeared to be no difference in the distribution of anti -- A and anti -- B activity in group O serum, though in two group O donors (J. F. and E. M.) only one type of agglutinin was found in the regions of low anionic binding capacity (Figs. 1 and 2).

Several samples of citrated plasma were fractionated in our laboratory by Method 6 of Cohn et al. These fractions were tested for ABO agglutinin activity, using fractions from group AB plasma as a control. As expected, most of the activity was found in Fraction ** f, with slight activity seen in Fraction 4, -- 1. A sample of Fraction ** f from group O plasma was dissolved in starting buffer, dialyzed against this buffer and subjected to chromatography using the gradient shown in Fig. 2. Once again, both anti -- A and anti -- B activities were found in the insoluble material precipitated during dialysis. Similarly, both types of antibodies were found in three regions of the chromatographic eluate, having extremely low, low, and high anionic binding capacity, respectively (Fig. 3).