Karyotype Analysis of Peripheral Blood
0.5mL peripheral blood of proband and her parents were cultured for 72
hrs at 37℃ with peripheral blood lymphocyte culture medium (Hangzhou
Biozone Science Technology). After that, 80 μL colchicine (working
concentration 10 μg/mL) was added and incubated for 65 min at 37℃.
Hypotonic treatment was performed with 8mL KCl solution (0.075 mol/L)
before 1mL fixed solution (methanol : glacial acetic acid = 3 : 1) was
used for fixation. Total 50 Giemsa–Trypsin–Giemsa (GTG)-banded
metaphase chromosomes were analyzed. For samples with abnormal
chromosomes, a total of 100 metaphases were analyzed.