Fig 2.

Comparisons of combined 45-min versus sequential (30 plus 45 min) incubations on killing of the antibody-coated pspC mutant, TRE118, in OPKA-2A and OPKA-2B. In the OPKA-2A, antibody-coated bacteria were incubated with BRC (complement) and HL-60 cells for 45 min. In OPKA-2B, the antibody-coated bacteria had separate sequential incubations with BRC for 30 min and HL-60 cells for 30 min. Rabbit serum specific for the family 1 PspA α-helical domain (A) and rabbit antiserum specific for the proline-rich domain containing a nonproline block (B) were diluted 1:10, 1:30, or 1:90 and evaluated for killing in OPKA-2A and OPKA-2B. A human serum sample J3 from a nonimmunized adult (C) was diluted 1:30, 1:100, and 1:300 and tested in OPKA-2A and OPKA-2B. In each panel, the percentages of the target bacteria killed in OPKA-2A are shown in black; the results for OPKA-2B are shown in red. The greater killing in OPKA-2B than in OPKA-2A was statistically significant at P values of 0.0009, 0.0004, and 0.035 for panels A, B, and C, respectively. The means and standard errors are shown for each data set. To calculate statistical significance between the treatments with each serum, we first ranked all 6 data points (three from the OPKA-2A and three from the OPKA-2B) at each dilution. Next (A and B), the ranked data points all for all three dilutions were grouped into two groups of 9 according to assay type. Next, the 9 data points for OPKA-2A were compared with the 9 for OPKA-2B by the Mann-Whitney two-sample rank test. With the human serum (C), the highest dilution showed no killing with either treatment. For this serum, only data from the 1:30 and 1:100 dilutions were used for statistical analysis.