Anaplastic Astrocytoma (Grade III) frozen primary human tissue obtained following surgical removal of astrocytic brain tumour. All samples used in this study were collected from brain tumour patients operated on at the neurosurgery departments of the Sahlgrenska Hospital, Göthenburg, Sweden or the Karolinska Hospital, Stockholm, Sweden. This study has been approved by the ethical committee of the Karolinska Hospital, Stockholm, Sweden. Tumour tissue was stored at -135C until use. Tumours were histopathologically diagnosed by Prof. V.P. Collins according to World Health Organization Brain tumour classification criteria. Total RNA was extracted using CsCl ultracentrifugation and asessed spectrophotometrically and using an Agilent Bioanalyzer 2100. Only RNA with no trace of degradation was used. 7 ug of total RNA were used for preparation of labelled target. Sample preparation, first and second strand cDNA synthesis, in vitro transcription amplification for production of biotin labelled cRNA, fragmentation of cRNA, U133A GeneChip hybridization, staining, washing and scanning were performed exactly as described by the manufacturer (Affymeytrix,Santa Clara). Hybridization was performed using 15 ug of labelled fragmented cRNA for 16 hours using a hybridization oven 640 as described by the manufacturer (Affymetrix, Santa Clara). Post-Hybridization washing and staining of the U133A GeneChips was performed on a fluidics station 400 (Affymetrix, Santa Clara) and scanning was performed on a GeneArray scanner (Affymetrix, Santa Clara).
