JMD ASIP MEMBERSHIP
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS

This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Purchase Article
Right arrow View Shopping Cart
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Matsukuma, S.
Right arrow Articles by Miyagi, Y.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Matsukuma, S.
Right arrow Articles by Miyagi, Y.
JMD 2006, Vol. 8, No. 4
Copyright © 2006 American Society for Investigative Pathology & Association for Molecular Pathology

Rapid and Simple Detection of Hot Spot Point Mutations of Epidermal Growth Factor Receptor, BRAF, and NRAS in Cancers Using the Loop-Hybrid Mobility Shift Assay

Shoichi Matsukuma*, Mitsuyo Yoshihara*, Fumio Kasai*, Akinori Kato*, Akira Yoshida{dagger}, Makoto Akaike{dagger}, Osamu Kobayashi{dagger}, Haruhiko Nakayama{dagger}, Yuji Sakuma*, Tsutomu Yoshida{ddagger}, Yoichi Kameda§, Eiju Tsuchiya* and Yohei Miyagi*{ddagger}

From the Division of Molecular Pathology and Genetics, * Kanagawa Cancer Center Research Institute, and the Division of Surgery, {dagger} the Laboratory for Molecular Diagnostics, {ddagger} and the Division of Pathology, § Kanagawa Cancer Center Hospital, Yokohama, Japan

A simple and rapid method to detect the epidermal growth factor receptor hot spot mutation L858R in lung adenocarcinoma was developed based on principles similar to the universal heteroduplex generator technology. A single-stranded oligonucleotide with an internal deletion was used to generate heteroduplexes (loop-hybrids) bearing a loop in the complementary strand derived from the polymerase chain reaction product of the normal or mutant allele. By placing deletion in the oligonucleotide adjacent to the mutational site, difference in electrophoretic mobility between loop-hybrids with normal and mutated DNA was distinguishable in a native polyacrylamide gel. The method was also modified to detect in-frame deletion mutations of epidermal growth factor receptor in lung adenocarcinomas. In addition, the method was adapted to detect hot spot mutations in the B-type Raf kinase (BRAF) at V600 and in a Ras-oncogene (NRAS) at Q61, the mutations commonly found in thyroid carcinomas. Our mutation detection system, designated the loop-hybrid mobility shift assay was sensitive enough to detect mutant DNA comprising 7.5% of the total DNA. As a simple and straightforward mutation detection technique, loop-hybrid mobility shift assay may be useful for the molecular diagnosis of certain types of clinical cancers. Other applications are also discussed.




This article has been cited by other articles:


Home page
 Clin. Cancer Res.Home page
W. Pao and M. Ladanyi
Epidermal Growth Factor Receptor Mutation Testing in Lung Cancer: Searching for the Ideal Method
Clin. Cancer Res., September 1, 2007; 13(17): 4954 - 4955.
[Full Text] [PDF]

Home page
 ChestHome page
T. Nakajima, K. Yasufuku, M. Suzuki, K. Hiroshima, R. Kubo, S. Mohammed, Y. Miyagi, S. Matsukuma, Y. Sekine, and T. Fujisawa
Assessment of Epidermal Growth Factor Receptor Mutation by Endobronchial Ultrasound-Guided Transbronchial Needle Aspiration
Chest, August 1, 2007; 132(2): 597 - 602.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Copyright © 2006 by the American Society for Investigative Pathology and the Association for Molecular Pathology.