An improved protocol of preparing bone marrow cells for fluorescence in situ hybridization / 中国实验血液学杂志
Journal of Experimental Hematology
;
(6): 496-499, 2012.
Artigo
em Chinês
| WPRIM
| ID: wpr-263364
ABSTRACT
This study was aimed to establish a smear protocol for preparing bone marrow cells and investigate its effect on fluorescence in situ hybridization (FISH) signal. Probe DNA (C-myc, MDM2, STK6) was labeled with Spectrum Green, PromoFluor-555 and PromoFluor-415 by nick translation. Five bone marrow samples were tested by two methods separately. Traditional method:
after removing the erythrocytes by hypoosmotic solution, the bone marrow cells were fixed in methanol/acetic acid (31). Improvedmethod:
erythrocytes were removed using density gradient centrifugation and fixed in methanol. The samples were then fixed again in 2 formaldehyde for 5 min. The FISH signal was assessed by comparing the relative signal intensity of each fluorophore with the autofluorescence background. The results indicated that improved method greatly increased the ratio of fluorescence signal intensity in the Spectrum Green, PromoFluor-555 and PromoFluor-415 channel (traditionalmethod:
4.3 ± 0.19, 3.52 ± 0.04, 3.07 ± 0.08; improvedmethod:
9.89 ± 0.41, 7.55 ± 0.5, 5.67 ± 0.18, n = 5, P < 0.01) respectively. The signal intensity increased 2.32, 2.14 and 1.85-fold in the Spectrum Green, PromoFluor-555 and PromoFluor-415 channel respectively. In addition, the improved method decreased the split signals [traditionalmethod:
(15.8 ± 1.74), (20.42 ± 2.88), (23.2 ± 3.02); improvedmethod:
(8.6 ± 1.2), (12.28 ± 1.33), (12.6 ± 2.56), n = 5, P < 0.05]. It is concluded that the improved optimal procedure which facilitates FISH intensity on bone marrow cells is developed, showing potential for wide application in the diagnosis of hematologic diseases.
Texto completo:
DisponíveL
Índice:
WPRIM (Pacífico Ocidental)
Assunto principal:
Células da Medula Óssea
/
Técnicas de Preparação Histocitológica
/
Hibridização in Situ Fluorescente
/
Biologia Celular
/
Métodos
Tipo de estudo:
Guia de Prática Clínica
Limite:
Humanos
Idioma:
Chinês
Revista:
Journal of Experimental Hematology
Ano de publicação:
2012
Tipo de documento:
Artigo
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