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Screen large numbers of novel drug compounds for their effect on the proliferation and erythroid-specific differentiation of human hematopoietic stem and progenitor cells (HSPCs).
Compared to using the colony-forming unit (CFU) assay, the liquid culture-based HemaTox™ assay is more flexible: you can add test compounds at various points throughout the culture and measure their effects on more primitive or more mature progenitor cells. The HemaTox™ Erythroid Kit promotes the proliferation of CD34+ HSPCs and their differentiation into cells expressing erythroid markers such as CD71 and CD235a.
Users can choose the readout method to quantify the response and estimate the IC50 and IC90 of each compound tested. Test up to 160 conditions—in triplicate—per kit in 5 x 96-well plates. This kit may be used on its own or in combination with HemaTox™ Myeloid Kit (Catalog #09704) or HemaTox™ Megakaryocyte Kit (Catalog #09707) to assess lineage-specific drug toxicity in parallel.
Subtype
Specialized Media
Cell Type
Erythroid Cells, Hematopoietic Stem and Progenitor Cells
Species
Human
Application
Cell Culture, Toxicity Assay
Brand
HemaTox
Area of Interest
Drug Discovery and Toxicity Testing, Stem Cell Biology
Figure 1. General HemaTox™ Erythroid Kit Procedure
*The cell isolation step may be omitted if pre-enriched CD34+ cells are used.
Figure 2. Flow Cytometry Plot Showing Erythroid Cells Produced After Culture of CD34+ HSPCs with the HemaTox™ Erythroid Kit
(A) Human CB CD34+ cells were cultured with the HemaTox™ Erythroid Kit using the protocol as written in the Product Information Sheet (PIS). (B) After the appropriate culture period, cells were harvested and stained for cell surface proteins expressed on erythroid (CD71 and GlyA) cells.
Figure 3. Reproducibility of HemaTox™ Erythroid Kit Results Between Experiments and Using Different CD34+ Cell Preparations
(A) Dose-response curves were generated from titrations of 5-FU added to human CB CD34+ cells isolated from 3 donors and cultured with the HemaTox™ Erythroid Kit. Three to five separate experiments were performed with cells from each donor. In each assay similar IC50 values were obtained with cells from different donors and in different experiments with cells from the same donor. Shown are values normalized to the percentages (%) of maximum cell growth without drug. Despite differences in absolute cell counts, curves are reproducible when normalized within each experiment. (B) Table showing IC50 values generated for 5-FU in culture with the erythroid kit including standard deviation (SD) and the coefficient of variation (% CV) across experiments.
Figure 4. Lineage-Specific Differences in Hematotoxicity Identified with HemaTox™ Erythroid, Myeloid and Megakaryocyte Kits
Results show average IC50 values for each drug tested on human BM CD34+ cells using the HemaTox™ Erythroid (grey), Myeloid (gold) and Megakaryocyte (orange) Kits. Most drugs show similar toxicity for each lineage but some, such as Sunitinib, are ~100-fold more toxic for erythroid than for megakaryocyte progenitor differentiation with intermediate toxicity for myeloid progenitor differentiation. Vertical lines indicate standard error of the mean (SEM) (n = 4 - 8).
Mouse monoclonal IgG1 antibody against human CD71 (transferrin receptor)
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HemaTox™ Erythroid Kit
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PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED. FOR ADDITIONAL INFORMATION ON QUALITY AT STEMCELL, REFER TO WWW.STEMCELL.COM/COMPLIANCE.