Reproducibility of SELDI Spectra Across Time and Laboratories

Abstract

This is an open access article. Unrestricted non-commercial use is permitted provided the original work is properly cited.The reproducibility of mass spectrometry (MS) data collected using surface enhanced laser desorption/ionization-time of flight (SELDI-TOF) has been questioned. This investigation was designed to test the reproducibility of SELDI data collected over time by multiple users and instruments. Five laboratories prepared arrays once every week for six weeks. Spectra were collected on separate instruments in the individual laboratories. Additionally, all of the arrays produced each week were rescanned on a single instrument in one laboratory. Lab-to-lab and array-to-array variability in alignment parameters were larger than the variability attributable to running samples during different weeks. The coefficient of variance (CV) in spectrum intensity ranged from 25% at baseline, to 80% in the matrix noise region, to about 50% during the exponential drop from the maximum matrix noise. Before normalization, the median CV of the peak heights was 72% and reduced to about 20% after normalization. Additionally, for the spectra from a common instrument, the CV ranged from 5% at baseline, to 50% in the matrix noise region, to 20% during the drop from the maximum matrix noise. Normalization reduced the variability in peak heights to about 18%. With proper processing methods, SELDI instruments produce spectra containing large numbers of reproducibly located peaks, with consistent heights.

Keywords: analysis of variance/reproducibility/SELDI; mass spectrometry/wavelet.

Figure 1.

A) Alignment offsets by ProteinChip. The arrays on the x-axis are ordered by week and by laboratory within week. Pairs of arrays from the same laboratory in the same week are adjacent. B) Alignment multiplier by ProteinChip. The arrays on the x-axis are ordered by week and by laboratory within week. Pairs of arrays from the same laboratory in the same week are adjacent.

Figure 2.

Reproducibility of spectrum intensity. These graphs contain plots of the mean intensity across 432 aligned spectra (top), the point-wise standard deviation (SD; center), and the point-wise coefficient of variation (CV; bottom) as functions of the time of flight.

Figure 3.

Results of a peak-by-peak decomposition of variance components before normalization. Each panel shows the percentage of variance of the log-transformed peak heights, as a function of the time-of-flight, for one of the factors (top left: residual; top right: array; lower left: laboratory; lower right: week).

Figure 4.

Variance of the log-transformed peak heights before and after normalization.

Figure 5.

Results of a peak-by-peak decomposition of variance components after normalization. Each panel shows the percentage of variance of the log-transformed peak heights, as a function of the time-of-flight, for one of the factors (top left: residual; top right: array; lower left: laboratory; lower right: week).