[BioC] OFF-TOPIC: Affy Scaling Factor and Absent/Present Calls

Laurent Gautier laurent at cbs.dtu.dk
Fri Apr 18 06:57:09 MEST 2003


On Fri, Apr 18, 2003 at 11:36:39AM +0800, Adaikalavan Ramasamy wrote:
> Dear all,
> 
> I have begun working with some affy data and there are some aspects of
> it that are confusing to me. The data was analysed using MAS 5.0 suite
> and most of the questions here pertain to single arrays (aka Absolute
> Analysis?).


Do you mean that you use the MAS software ? If this is the case,
we might be in the same position as you (i.e. with the official technical
documentation provided, especially the sadd_white-paper.pdf that
can be found on the website of the company). Did you read this document ?


Regards,


L.


> 
> The first 3 question is on Scaling Factors (SF) which is defined as 
> 
> 	SF = TGT / x_96 	; where x_96 is the 2% trimmed mean 
> 
> 
> 1. Why does MAS 5.0 apply the calculated SF to the middle 96% only and
> not 100% of the data ? Since it is often that SF > 1 and signal > 0 by
> affy's definition, the top 2% are being penalized. What is the rational
> behind this ?
> 
> 2. I have read that a SF > 3 generally implies problems with the
> experiment. Why is this?
> 
> 3. How is the target intensity, TGT, chosen ? Values of 100, 200, 500
> and even 1500 seem to have been chosen. Or do we choose TGT to ensure SF
> < 3 for most of the arrays.
> 
> 
> The next 2 are related to the Present/Absent calls provided.
> 
> 4. I have 74 arrays and on average 50% of the data is missing. What is
> the normal/acceptable observed percentage missing.
> 
> 5. I am removing all the signal data with Absent Call. Is this correct
> and is there any other use for Absent Call.
> 
> 
> Googling and reading the affy manuals (some parts of it anyway) has not
> cleared these confusions. Any answers, advices or references from this
> list is much appreciated. Thank you for your time.
> 
> Regards, Adai.
> 
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