Assessment of DNA Viability in Long Term-Stored Buffy Coat Species for the Korean Multicenter Cancer Cohort

Yang, Mihi; Yoo, Jihyun; Kim, Cheong Sik; Shin, Aesun; Kang, Daehee; Chang, Soung Hoon; Park, Sue Kyung; Shin, Hai Rim; Yoo, Keun Young

Abstract

OBJECTIVES
Peripheral blood-buffy coat fractions (N = 14, 956) have been stored at -70degrees C in the headquarter of the Korean Multicenter Cancer Cohort (KMCC), since 1993. To study the future molecular etiology of cancers using specimens of the cohort, properly stored specimens are necessary. Therefore, the DNA-viability of the buffy coat samples was investigated. METHODS: Buffy coat fraction samples were randomly selected from various collection areas and years (N = 100). The DNA viability was evaluate from the UV-absorbent ratios at 260/280nm and the PCR for beta-globin was performed with genomic DNA isolated from the buffy coat. RESULTS: PCR products were obtained from 85 and 98% of the C and H area-samples, respectively, using 50 or 100mul of the buffy coat. There were significant differences in the yields of the PCR-amplifications from the C and H areas (p < 0.05), which was due to differences in the homogenization of the buffy coat fractions available as aliquots. The PCR-products were obtained from all of the samples (N = 7) stored at the C area-local center, but the other aliquots stored at the headquarter were not PCR-amplified. Therefore, the PCR products in almost all the samples, even including the DNA-degraded samples, were obtained. In addition, an improvement in the DNA isolation, i.e. approx. 1.6 fold, was found after using extra RBC lysis buffer. CONCLUSIONS: PCR products for beta-globin were obtained from nearly all of the samples. The regional differences in the PCR amplifications were thought to have originated from the different sample-preparation and homogenization performance. Therefore, the long term-stored buffy coat species at the KMCC can be used for future molecular studies.