[R-sig-ME] Mixed Models - remedy for pseudoreplication?

[Jens Oldeland]

Dear List readers,

we are studying effects of chemical therapies on the proliferation of 
cancer cell lines.
My question aims at the practical application of a Mixed Effect Model 
(lme in R) of a particular analysis of a cell culture experiment on a 96 
well plate and the problem of pseudoreplication.

A 96 well plate looks like this: 
http://www.cellsignet.com/media/plates/96.jpg
and receives treatments in the columns 3-10 with 8 wells, i.e. the 
replications. We repeated the experiment on 6 days, with one 96 well 
plate per day.
We noticed a high intra-day variability.

Since all the cells of one day on which the treatments are applied come 
from the same flask and the media and pharmacologic agents, with which 
they were treated, are coming from the same tubes, the question arises 
if I have to count them as pseudoreplicates.
The analysis of the data using “lme” works just fine, but I worry about 
the high number of degrees of freedom, which include the 
“pseudoreplicated” measurements.

An alternative would be to average the 8 replicates per day and use an 
ANOVA with an Error structure, which seems less elegant to me, since we 
are not including the high intra-day variability.

I would be very happy if anyone could give an adivce on how to deal with 
this dataset. I found different opinions in the literature, some saying 
that LMMs are a "remedy" for 96 well plates and some say that 96 well 
plates are per-se pseudoreplicated...

thanks in advance!
Jens

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Dr. Jens Oldeland

Post-Doc Researcher & Lecturer @ BEE
Managing Editor - Biodiversity & Ecology

Biodiversity, Ecology and Evolution of Plants (BEE)
Biocentre Klein Flottbek and Botanical Garden
University of Hamburg
Ohnhorststr. 18
22609 Hamburg,
Germany

Tel:    0049-(0)40-42816-407
Fax:    0049-(0)40-42816-543
Mail: 	jens.oldeland at uni-hamburg.de
         Oldeland at gmx.de 	
Skype:	jens.oldeland
http://www.biologie.uni-hamburg.de/bzf/fbda005/fbda005.htm
http://www.biodiversity-plants.de/biodivers_ecol/biodivers_ecol.php
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