A hanging drop culture method to study terminal erythroid differentiation.

Abstract:

OBJECTIVE:To design a culture method allowing the quantitative and qualitative analysis of terminal erythroid differentiation. METHODS:Primary erythroid progenitors derived either from mouse tissues or from human umbilical cord blood were differentiated using hanging drop cultures and compared to methylcellulose cultures. Cultured cells were analyzed by FACS to assess differentiation. RESULTS:We describe a practical culture method by adapting the previously described hanging drop culture system to conditions allowing terminal differentiation of primary erythroid progenitors. Using minimal volumes of media and small numbers of cells, we obtained quantitative terminal erythroid differentiation within two days of culture in the case of murine cells and 4 days in the case of human cells. CONCLUSIONS:The established methods for ex vivo culture of primary erythroid progenitors, such as methylcellulose-based burst-forming unit-erythroid (BFU-E) and colony-forming unit-erythroid (CFU-E) assays, allow the detection of committed erythroid progenitors but are of limited value to study terminal erythroid differentiation. We show that the application of hanging drop cultures is a practical alternative that, in combination with clonogenic assays, enables a comprehensive assessment of the behavior of primary erythroid cells ex vivo in the context of genetic and drug-induced perturbations.

journal_name

Exp Hematol

journal_title

Experimental hematology

authors

Gutiérrez L,Lindeboom F,Ferreira R,Drissen R,Grosveld F,Whyatt D,Philipsen S

doi

10.1016/j.exphem.2005.06.014

keywords:

subject

Has Abstract

pub_date

2005-10-01 00:00:00

pages

1083-91

issue

10

eissn

0301-472X

issn

1873-2399

pii

S0301-472X(05)00282-1

journal_volume

33

pub_type

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