Figure 6

Affymetrix spiked-in experiment: The upper panel shows the hook obtained from one chip of this series. The predominant number of probes is hybridized with RNA of a HeLa-cell extract which was added to the chips to mimic a complex hybridization background (thick blue curve). The spike-probe sets are indicated by the open symbols and the respective transcript concentrations (see the numbers, the concentrations are given in units of pM). The horizontal distance between a spike position and the end point is related to the logarithm of the specific binding strength. The turning point between the N- and the mix-ranges defines the threshold for present probes. The dashed line is the fit of the Langmuir hybridization model to the data. The middle and lower parts show present/absent characteristics and the S/N-ratio of the spikes, respectively. The fraction of absent probes and the S/N ratio were calculated as mean values over all 42 chips of the experimental series (see thick lines). The open circles in the lower part show the individual probe-set values and thus the scatter of these points about their mean value. Spiked probes with nominal concentrations larger than 2 pM are "safely" called present. The S/N-ratio linearly correlates with the spiked-in concentration. The right axis of the lower part scales the expression estimates in units of the binding strength. The green dashed lines indicated that the threshold for calling probes as present corresponds to S/N-ratios R ≈ 0.1 – 2 and the S-binding strength of X^N ≈ (0.5 – 5) 10^-3.