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Table 3 Kinetic parameters obtained from the optimization of the Peptidomatrix conditions with the peptides and antibodies specific to Na/K ATPase (see figure 4) (± SD, n = 3)

From: Competition between bound and free peptides in an ELISA-based procedure that assays peptides derived from protein digests

A: increasing attached peptide at two fixed levels of added antibody

 

1 stAB Concentration 0.00033 serum/PBS

1 stAB Concentration 0.00066 serum/PBS

 

Kd (μg/ml)

0.19 ± 0.01

0.1 ± 0.02

 

Amax (OD units)

1.36 ± 0.02

1.56 ± 0.08

 

D (OD units)

0.25 ± 0.02

0.38 ± 0.07

 

B: Competition assay – Kd determined when increasing added free peptide at three fixed levels of added antibody

 

1 stAB concentration 0.000625 serum/PBS

1 stAB concentration 0.000125serum/PBS

1 stAB concentration 0.0025 serum/PBS

Kd (μg/ml)

0.55 ± 0.1

0.83 ± 0.12

2.55 ± 1.21

Amax (OD units)

1.16 ± 0.05

1.35 ± 0.05

1.33 ± 0.17

D (OD units)

0.59 ± 0.04

0.86 ± 0.04

1.17 ± 0.18

C: increasing attached peptide at two fixed levels of added antibody

 

1 stAB concentration 0.00033 serum/PBS

1 stAB concentration 0.00067 serum/PBS

 

Kd (serum dilutions)

0.33 ± 0.08

0.29 ± 0.04

 

Amax (OD units)

1.46 ± 0.09

1.74 ± 0.06

 

D (OD units)

0.19 ± 0.079

0.24 ± 0.05

 

D: Competition assay- Kd determined when increasing added free peptide at three fixed levels of added antibody

 

1 stAB concentration 0.000625 serum/PBS

1 stAB concentration 0.00125 serum/PBS

1 stAB concentration 0.0025 serum/PBS

Kd (μg/ml)

0.68 ± 0.12

0.99 ± 0.19

2.04 ± 0.3

Amax (OD units)

1.58 ± 0.06

1.87 ± 0.09

2.05 ± 0.08

D (OD units)

0.25 ± 0.05

0.25 ± 0.08

0.28 ± 0.08