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

Adaikalavan Ramasamy gisar at nus.edu.sg
Tue Apr 22 19:36:37 MEST 2003

Dear Laurent, 

Thank you for the reference. I have just finishing reading it and I
think I am more confused because on page 8, it appears that they
intended to multiply the scaling factor to 100% of the data. 

" The reported value of probe set i is: ReportedValue(i) = nf * sf * 2 ^
SignalLogValue "

My data is contrary to this. If possible, could you check the mean value
and the 2% trimmed mean after applying to scale factor to eliminate the
possibility error on my part. Much appreciated.

Can anyone shed some light on the remaining questions ? Or is there an
affy mailing list group I can ask. Thanks.

Regards, Adai.

-----Original Message-----
From: Laurent Gautier [mailto:laurent at cbs.dtu.dk] 
Sent: Friday, April 18, 2003 11:57 AM
To: Adaikalavan Ramasamy
Cc: Bioconductor mailing list
Subject: Re: [BioC] OFF-TOPIC: Affy Scaling Factor and Absent/Present

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 ?



> 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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