Dear Hans-Ulrich et al., vsn >= 3.4.11. now computes separate offsets for different strate (such as e.g. print-tip groups), as suggested by Hans-Ulrich earlier in this thread. It is available at http://www.bioconductor.org/packages/2.2/bioc/html/vsn.html Best wishes Wolfgang ------------------------------------------------------------------ Wolfgang Huber EBI/EMBL Cambridge UK http://www.ebi.ac.uk/huber 04/03/2008 16:52 Wolfgang Huber a ?crit > Dear Hans-Ulrich, > > thank you for your thoughtful message! The executive summary: this is > indeed an (unintended) difference between vsn and vsn2, and I will > update vsn2 before the next release. It only affects applications with > multiple strata (print-tip groups). > > Bacgkround: the error and normalisation model of vsn is invariant under > an overall scaling of the data: if you multiply all intensities by a > factor of 10, you will get the same output - except for an overall shift > on the glog2 scale of log2(10). This makes sense because microarray data > don't have units and a value of "200" can mean very different things say > on an Affymetrix genechip and on a custom-made array. > > This explains why there is this 'arbitrary' offset c. It is computed > through an explicite formula from the b's (i.e. the scale factors), > hence the fact whether your actual data contain instances of large x > does not directly matter (it may indirectly, by affecting how the b's > are estimated). For x -> infinity, the function glog2(f(b)*x+a) > approaches log2(x) + log2(f(b)) + log2(2), and c is computed to cancel > out the last two terms, so that for large x, the net transformation > resembles log2(x). There is one b for each array and stratum (=print tip > group). The current implementation of vsn2 computes one single value c > by taking the mean of log2(f(b)) + log2(2) across all strata and arrays. > The old vsn computed c from the b's of the first array only, but > separately for each stratum. > > I had not anticipated that the difference between strata could make such > a difference, but given your observations, and with more thought about > it, it does make sense. I will update vsn2 to compute c from averaging > over the arrays, but separately for each stratum. > > Best wishes > Wolfgang > > ------------------------------------------------------------------ > Wolfgang Huber EBI/EMBL Cambridge UK http://www.ebi.ac.uk/huber > > > 04/03/2008 16:38 Hans-Ulrich Klein scripsit >> Dear all, >> >> I use the vsn2 method to normalize single-colour arrays with 48 >> print-tips (25*26 oligos per print-tip). After normalization, the >> intensities of the 48 print-tips are in different ranges. The grid of >> the print-tips can be seen clearly on false color representations of the >> arrays' spatial distributions of feature intensities. However, scale and >> location of the intensities of a print-tip do not change across arrays. >> >> The man page of vsn2 says: >> "The data are returned on a glog scale to base 2. More precisely, >> the transformed data are subject to the transformation >> glog2(f(b)*x+a) + c, where glog2(u) = log2(u+sqrt(u*u+1)) = >> asinh(u)/log(2) is called the generalised logarithm, a and b are >> the fitted model parameters (see references), f is a parameter >> transformation [4], and the overall constant offset c is computed >> from b such that for large x the transformation approximately >> corresponds to the log2 function." >> >> May be there are not enough "large x" in some print-tips due to missing >> values in my data. I observed that reducing the number of oligos leads >> to even larger differences in the print-tip offsets. Are there >> parameters to take influence on the computation of c? Has someone else >> observed this problem? The older "vsn" function does not lead to >> different print-tip offsets. >> >> Regards, >> Hans-Ulrich >> >> >> >> >> > sessionInfo() >> R version 2.6.2 (2008-02-08) >> x86_64-pc-linux-gnu >> >> locale: >> C >> >> attached base packages: >> [1] tools stats graphics grDevices utils datasets methods >> [8] base >> >> other attached packages: >> [1] vsn_3.2.1 limma_2.12.0 affy_1.16.0 >> [4] preprocessCore_1.0.0 affyio_1.6.1 Biobase_1.16.3 >> >> loaded via a namespace (and not attached): >> [1] grid_2.6.2 lattice_0.17-4 rcompgen_0.1-17 >> >> _______________________________________________ >> Bioconductor mailing list >> Bioconductor at stat.math.ethz.ch >> https://stat.ethz.ch/mailman/listinfo/bioconductor >> Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor > > _______________________________________________ > Bioconductor mailing list > Bioconductor at stat.math.ethz.ch > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor