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Molecular pathology detection

LK Ling-Wei Kong
JC Jin Chen
HZ Heng Zhao
KY Kun Yao
SF Sheng-Yu Fang
ZW Zheng Wang
YW Yin-Yan Wang
SL Shou-Wei Li
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IDH1 mutation proteins were detected by immunohistochemistry, and proteins showing negative expression were used to determine the IDH1 mutation status by using DNA pyrosequencing28, First, complete genomic DNA was extracted from tumor tissue samples using a QIAamp DNA Mini Kit (QIAGEN, Hilden, Germany). Second, the genomic region spanning the Wild-type R132 of IDH1 was amplified with the primers 5′-GCTTGTGAGTGGATGGGTAAAAC-3′ and 5′-biotin-TTGCCAACATGACTTACTTGATC-3′. Then, the MGMT methylation status was determined by pyrosequencing using a 5′- GGATATGTTGGGATAGT-3′ primer aimed at the MGMT promoter region. A total of seven methylation sites were detected, and a methylation percentage greater than 10% was considered to indicate hypermethylation. Finally, fluorescence in situ hybridization was used to identify 1p19q deletion status.

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