. 2015 Aug 7;4:e07467. doi: 10.7554/eLife.07467

Table 2.

IC₇₅ values for anti-PR8 lamprey plasmas and guinea pig serum (positive control) in competition with defined HA mAbs by ELISA^*

DOI: http://dx.doi.org/10.7554/eLife.07467.014

Table 2—source data 1.

Competition ELISA against α-Head HA panel Abs. Data from Table 2 shown in graph form. Serially diluted unlabeled lamprey plasma raised against PR8 (L7, L9 or Naïve) was added to PR8 immobilized on 96 well ELISA plates. After 1 hr incubation, a fixed concentration of each indicated hybridoma supernatant (PEG-1, H28E23, H18 S413, H35 C12, H2 4B1, H18 S210, and Y8 2D1) was added at a predetermined concentration—65% of maximum binding (EC65). Data from three independent experiments were analyzed by Two Way ANOVA followed by Bonferroni Multiple Comparisons against the Naïve plasma data using PRISM. (*p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001).

DOI: http://dx.doi.org/10.7554/eLife.07467.015

elife07467s004.pdf^{(2.1MB, pdf)}

DOI: 10.7554/eLife.07467.015

Table 2—source data 2.

Competition ELISA against α-Head HA Fabs. Same as Table 2—source data 1 but with Fabs instead of hybridoma supernatants. p-value measurements determined with One-Way Anova followed by Dunnett's Multiple Comparison Test against Naïve plasma values. Stars indicate differences among whole groups. Data collected from only one experiment due to shortage of lamprey plasma.

DOI: http://dx.doi.org/10.7554/eLife.07467.016

elife07467s005.pdf^{(1.9MB, pdf)}

DOI: 10.7554/eLife.07467.016

Table 2—source data 3.

Immune lamprey plasma does not compete against stem binding Abs by ELISA. Serially diluted naïve or immune lamprey plasma raised against PR8 (L29) on 96 well ELISA plates immobilized with PR8. After 1 hr incubation, a fixed concentration of purified monoclonal C179 or 2G02 was added at EC65. As a positive control, the two stem Abs were competed against each other or against an anti-HA head Ab (H28E23). Data are from at least two separate experiments with four total replicates. There was no statistical difference between the lamprey plasma curves. ELISA signal from these Abs is low, thus the curves are noisy. In contrast, the ‘2G02 then C179 curve’ is statistically different from the ‘H28E23 then 2G02 curve’ by two-tailed t-test (**p< 0.01).

DOI: http://dx.doi.org/10.7554/eLife.07467.017

elife07467s006.pdf^{(403.7KB, pdf)}

DOI: 10.7554/eLife.07467.017

	L7		L9		L29		Guinea pig	Stem
Epitope^†	IgG	Fab	IgG	Fab	IgG	Fab	IgG	C179
Sa	NC^‡	NC	250	400	—	290	36,000	NC
Sb	NC	NC	320	410	—	530	17,000	—
Cb	280	610	600	980	—	860	26,000	—
Ca1	250	350	360	300	—	860	40,000	—
Ca2	370	—	600	—	—	—	25,000	—
Stem	—	—	—	—	NC	—	—	3.5
2G02	–	–	–	–	–	–	–	nM

Data was fit to a Hill Slope. IC₇₅ value was calculated from the curve using PRISM. IgG data are from three independent experiments, Fab data are from one experiment due to limited lamprey plasma.

^†

mAbs used—Sa: PEG-1; Sb: H28E23; Cb: H36 C12 (IgG), H9 D3 (Fab); Ca1: H2 4B1; Ca2: H18 S413; Stem: C179 and 2G02.

^‡

NC, no competition; ‘—’, not determined.

mAbs, monoclonal antibodies.