Naslov
The impact of number of leukocytes on concentration and purity of isolated DNA

Autori
Đuraković, Melani ; Rumora, Lada ; Ćelap, Ivana ; Šutić, Maja ; Grdić Rajković, Marija

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Biochemia Medica / Pašalić, Daria - Zagreb : Hrvatsko društvo za medicinsku biokemiju i laboratorijsku medicinu (HDMBLM), 2018, S133-S134

Skup
9. kongres hrvatskog društva za medicinsku biokemiju i laboratorijsku medicinu (HDMBLM)

Mjesto i datum
Zagreb, Hrvatska, 09.05.2018. - 12.05.2018

Vrsta sudjelovanja
Poster

Vrsta recenzije
Domaća recenzija

Ključne riječi
DNA : isolation ; leukocytes

Sažetak
Isolation of DNA is a key step that precede various analysis in molecular diagnostics. Extracting DNA from leukocytes by salting out method (by Miller or with Qiagen purification kit) is based on increasing salt concentration which separates proteins and cellular debris from the DNA fraction. After centrifugation of the mixture, DNA molecules are precipitated by absolute ethanol. Purpose of this research is to examine the impact of number of leukocytes on concentration and purity of isolated DNA. Number of leukocytes was determined on automated hematology analyser immediately after sampling of blood. Whole blood samples taken on anticoagulant EDTA were stored on -20 °C until DNA extration. DNA was isolated by Miller method (N=227 ; 92 healthy subjects and 135 subjects with chronic obstructive pulmonry disease ) and Qiagen commercial kit (N=173 ; 98 healthy subjects and 75 patients undertaking hemodialysis). Apsorbances at 260 and 280 were measured on DeNovix DS-11 highly sensitive spectrophotometer. Statistic analysis was done by MedCalc statistic program. Higher DNA concentrations were isolated by Miller method compared to the Qiagen comercial kit method [138.6 (123.1-151.7) vs 57.0 (48.0-76.5) ng/µL ; P<0.001 and purity [1.7 (1.7-1.7) vs 1.3 (1.3- 1.5) ; P<0.001]. Number of leukocytes in samples used for two methods of isolation was significantly different. We observe weak correlation between number of leukocytes and DNA concentration (r=0.27 ; P<0.001 and r=0.43 ; P<0.001) and between number of leukocytes and DNA purity (r=0.38 ; P<0.001 and r=0.27 ; P<0.001). Depending on method, number of leukocytes showed significant but weak impact on extracted DNA concentration and purity. However, it is very important to know that before isolation all samples were frozen, which could have an effect on the obtained results.

Izvorni jezik
Engleski

Znanstvena područja
Temeljne medicinske znanosti, Kliničke medicinske znanosti, Farmacija

POVEZANOST RADA