Table 3. Pure Mast Cell Colony Formation... | American Society of Hematology

Table 3.

Pure Mast Cell Colony Formation Derived From CD34⁺ Cells in the Presence of SF, IL-6, and IL-3 (With or Without IL-4)

(Cytokines) Phenotypes	Plated Cells	Mixed Type Colonies^3-150	Pure Cell Type Colonies^3-151	Pure Mast Cell Colonies^3-152
(SF, IL-6, IL-3)
CD34⁺CD38⁺	600	12	40	11/16
CD34⁺CD38⁻	600	194^3-153	14	2/9

(SF, IL-6, IL-3)
CD34⁺HLA-DR⁺	600	115^3-153	16	10/13
CD34⁺HLA-DR⁻	600	140^3-153	39	23/30

(SF, IL-6, IL-3, IL-4)
CD34⁺HLA-DR⁺	600	26^3-155	7	1/7
CD34⁺HLA-DR⁻	600	23^3-154	13	6/8

(Cytokines) Phenotypes	Plated Cells	Mixed Type Colonies^3-150	Pure Cell Type Colonies^3-151	Pure Mast Cell Colonies^3-152
(SF, IL-6, IL-3)
CD34⁺CD38⁺	600	12	40	11/16
CD34⁺CD38⁻	600	194^3-153	14	2/9

(SF, IL-6, IL-3)
CD34⁺HLA-DR⁺	600	115^3-153	16	10/13
CD34⁺HLA-DR⁻	600	140^3-153	39	23/30

(SF, IL-6, IL-3, IL-4)
CD34⁺HLA-DR⁺	600	26^3-155	7	1/7
CD34⁺HLA-DR⁻	600	23^3-154	13	6/8

CD34⁺ cells were purified using Macs II (Miltenyl Biotec). After magnetic microbeads being removed with the CD34-isolation kit, the cells were further separated with either for CD38 or HLA-DR. The 300 cells of each fraction were seeded in 1 mL methylcellulose (0.9%) supplemented with 100 ng/mL SF, 50 ng/mL IL-6, and 5 ng/mL IL-3 (with or without 10 ng/mL IL-4. On day 14 in culture, 1 mL methylcellulose (0.6%) supplemented with SF and IL-6 (without cells) was further layered over the methylcellulose culture. They were cultured for a total of 28 days. The experiment was repeated twice, and the total number of colonies was shown. A colony was lifted, divided into two parts, and stained with May-Giemsa or antitryptase MoAb.

F3-150

Colonies consisting of heterogeneous sized cells.

F3-151

Colonies consisting of homogenous sized cells. Colonies consisting of macrophages and of degenerated cells were excluded for the counting.

F3-152

Pure mast cell colony was defined as follows: (1) A colony consisted of mast-like cells or immature cells but not any other lineages, including basophil-like cells. (2) In addition, the colony should consist of greater than 50% tryptase-positive cells. The colonies are expressed as the number of colonies examined with staining out of the number of pure cell type colonies. Pure cell type colonies without mast cells consisted mainly of eosinophils or immature monocytic cells.

F3-153

Mast cells were found at 37 of 40 mixed cell colonies examined with staining.

F3-155

Mast cells were found at 8 of 20 mixed cell colonies examined with staining.

F3-154

Mast cells were found at 2 of 6 mixed cell colonies examined with staining.

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