Portions the size of 1/6 to 1/32 part of unincubated blastoderm cultured in an albumen-salineagar solid medium form erythroid cells under conditions which suppress normal co-ordinated movements for mesoderm induction. Anterior and posterior regions of unincubated blastoderm have the same potentiality to form erythroid cells. The marginal zone seems to be the contributor of prospective erythroid cells. Portion 1/16 part of unincubated blastoderm forms morphologically distinct erythroid cells of the primitive and definitive lines as in normal development in ovo. It seems progenitor cell(s) is preprogrammed to a particular pattern of differentiation and/or includes differentiation to various erythroid cell types as an obligatory step. This system provides novel experimental possibilities in the study of erythroid cell determination and differentiation.The blastoderm of the newly laid chicken egg contains about 60,000 cells and consists of the 1 to 2-cell thick area pellucida and the thicker area opaca (1). The different regions of the pre-streak blastoderm are not completely equivalent to blood cell formation, but there is a definite region, more or less firmly commited to erythroid cell differentiation, which will give rise to the area opaca vasculosa (a. 0. v.) (2). However, this "determined" state can be shifted and multiple blood forming areas can be induced in the pre-streak blastoderm (3). It has been reported that stem cell(s), which give rise to primitive and definitive erythroid cell lines, arise in the yolk sac outside the embryonic body (4, 5, 6). Unincubated blastoderm in which the primitive streak (PS) and, consequently, the embryonic body are inhibited mechanically can form primitive and definitive erythrocytes and embryonic and adult hemoglobins (Hb) as in development in o w (6).In the work presented in this report, we determined the critical mass of portions from unincubated blastoderm capable of forming erythroid cells under conditions which suppress normal tissue associations and tissue interactions. In addition, we studied their capacity of forming erythroid cells characteristic of the primitive and definitive lines. These results give insights concerning the regulation of erythroid cell determination and differentiation in the early chick blastoderm.
MATERIALS AND METHODS
CultureFreshly laid White Leghorn chicken fertilized eggs were used. Unincubated blastoderms (stage X) (7) were removed from the yolk, washed free of the vitelline membrane and most of the adhering yolk, and carried in a drop of phosphate buffered saline solution (PBS-DULBECCO'S) with a widemouthed medicine dropper on SPRATT'S albumen-saline-agar (we substituted plain saline by PBS) solid medium.Blastoderms were stretched and dissected into 4, 8, 14, 16, 32 and smaller V (wedge) -shaped equal portions, their apices coincident with the center of the blastoderm, with fine dissecting steel needles. In some cases, portion 95