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H4034

Manufactured by STEMCELL
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Sourced in Canada, France

H4034 is a laboratory equipment product manufactured by STEMCELL. It is designed for cell culture applications.

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Lab products found in correlation

Trypsin edta, by Thermo Fisher Scientific (3 mentions) Stemvision, by STEMCELL (2 mentions) L glutamine, by Miltenyi Biotec (1 mentions) Alpha mem, by Thermo Fisher Scientific (1 mentions) Thrombopoietin tpo, by Miltenyi Biotec (1 mentions) Penicillin streptomycin, by Miltenyi Biotec (1 mentions) Ficoll paque, by Thermo Fisher Scientific (1 mentions) Tgf β, by Thermo Fisher Scientific (1 mentions) Anti mouse igg1 microbeads, by Miltenyi Biotec (1 mentions) Methocult gf m3434, by STEMCELL (1 mentions) G csf, by Thermo Fisher Scientific (1 mentions) Gm csf, by Thermo Fisher Scientific (1 mentions) Erythropoietin epo, by Thermo Fisher Scientific (1 mentions) Virapower, by Thermo Fisher Scientific (1 mentions) Inverted microscope, by Nikon (1 mentions)

15 protocols using h4034

1

hESC-derived Colony Forming Assays

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Day 5 hESC-derived cells were harvested as single cells and plated for colony forming-unit (CFU) progenitors in methylcellulose containing human cytokines (H4034, Stem Cell Technologies). Colonies were scored by morphology after 12–14 days as small, primitive erythroid (CFU-EryP), and macrophage (CFU-Mac).
Palpant N.J., Wang Y., Hadland B., Zaunbrecher R.J., Redd M., Jones D., Pabon L., Jain R., Epstein J., Ruzzo W.L., Zheng Y., Bernstein I., Margolin A, & Murry C.E. (2017). Chromatin and transcriptional analysis of mesoderm progenitor cells identifies HOPX as a regulator of primitive hematopoiesis. Cell reports, 20(7), 1597-1608.
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2

Quantifying Hematopoietic Stem Cell Potency

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3 days post RNP-electroporation, 500 CD34+ HSPCs were plated in 1ml methylcellulose media (# H4034, Stem Cell Technologies). Primary CFU-C colonies were counted after 14 days. For the colony replating experiments, 2 weeks after the primary plating, the colonies from three pates were pooled, washed with PBS, and the cells were plated in new methylcellulose media at 25,000 cells/ml for an additional 2 weeks.
Bick A.G., Weinstock J.S., Nandakumar S.K., Fulco C.P., Bao E.L., Zekavat S.M., Szeto M.D., Liao X., Leventhal M.J., Nasser J., Chang K., Laurie C., Burugula B.B., Gibson C.J., Lin A.E., Taub M.A., Aguet F., Ardlie K., Mitchell B.D., Barnes K.C., Moscati A., Fornage M., Redline S., Psaty B.M., Silverman E.K., Weiss S.T., Palmer N.D., Vasan R.S., Burchard E.G., Kardia S.L., He J., Kaplan R.C., Smith N.L., Arnett D.K., Schwartz D.A., Correa A., de Andrade M., Guo X., Konkle B.A., Custer B., Peralta J.M., Gui H., Meyers D.A., McGarvey S.T., Chen I.Y., Shoemaker M.B., Peyser P.A., Broome J.G., Gogarten S.M., Wang F.F., Wong Q., Montasser M.E., Daya M., Kenny E.E., North K.E., Launer L.J., Cade B.E., Bis J.C., Cho M.H., Lasky-Su J., Bowden D.W., Cupples L.A., Mak A.C., Becker L.C., Smith J.A., Kelly T.N., Aslibekyan S., Heckbert S.R., Tiwari H.K., Yang I.V., Heit J.A., Lubitz S., Johnsen J.M., Curran J.E., Wenzel S.E., Weeks D.E., Rao D.C., Darbar D., Moon J.Y., Tracy R.P., Buth E.J., Rafaels N., Loos R.J., Durda P., Liu Y., Hou L., Lee J., Kachroo P., Freedman B.I., Levy D., Bielak L.F., Hixson J.E., Floyd J.S., Whitsel E.A., Ellinor P.T., Irvin M.R., Fingerlin T.E., Raffield L.M., Armasu S.M., Wheeler M.M., Sabino E.C., Blangero J., Williams L.K., Levy B.D., Huey-Herng Sheu W., Roden D.M., Boerwinkle E., Manson J.E., Mathias R.A., Desai P., Taylor K.D., Johnson A.D., Auer P.L., Kooperberg C., Laurie C.C., Blackwell T.W., Smith A.V., Zhao H., Lange E., Lange L., Rich S.S., Rotter J.I., Wilson J.G., Scheet P., Kitzman J.O., Lander E.S., Engreitz J.M., Ebert B.L., Reiner A.P., Jaiswal S., Abecasis G., Sankaran V.G., Kathiresan S, & Natarajan P. (2020). Inherited Causes of Clonal Hematopoiesis in 97,691 TOPMed Whole Genomes. Nature, 586(7831), 763-768.
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3

Hematopoietic Stem Cell Assay

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Cells were pre-treated with GDYO or GO (20 μg/mL) for 12 h. 1,000 cells were seeded in methylcellulose media (H4034, Stemcell Technologies, Inc.) in triplicates. After 10 days, plates were scored for colony number on microscopy.
Wang Q., Liu Y., Wang H., Jiang P., Qian W., You M., Han Y., Zeng X., Li J., Lu H., Jiang L., Zhu M., Li S., Huang K., Tang M., Wang X., Yan L., Xiong Z., Shi X., Bai G., Liu H., Li Y., Zhao Y., Chen C, & Qian P. (2022). Graphdiyne oxide nanosheets display selective anti-leukemia efficacy against DNMT3A-mutant AML cells. Nature Communications, 13, 5657.
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4

Evaluating HSCs Progenitor Potential

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The evaluation of HSCs progenitor potential in treated groups bone marrow c-Kit+ cells were sorted and seeded in methylcellulose medium (#H4034, Stem Cell Technologies) at a concentration of 1 × 103 cells /ml in triplicates. Numbers of colonies were counted after 7 days and scored by standard morphologic criteria.
Yadav M., Song F., Huang J., Chakravarti A, & Jacob N.K. (2018). Ocimum flavone Orientin as a countermeasure for thrombocytopenia. Scientific Reports, 8, 5075.
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5

Quantifying Hematopoietic Stem Cell Potency

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3 days post RNP-electroporation, 500 CD34+ HSPCs were plated in 1ml methylcellulose media (# H4034, Stem Cell Technologies). Primary CFU-C colonies were counted after 14 days. For the colony replating experiments, 2 weeks after the primary plating, the colonies from three pates were pooled, washed with PBS, and the cells were plated in new methylcellulose media at 25,000 cells/ml for an additional 2 weeks.
Bick A.G., Weinstock J.S., Nandakumar S.K., Fulco C.P., Bao E.L., Zekavat S.M., Szeto M.D., Liao X., Leventhal M.J., Nasser J., Chang K., Laurie C., Burugula B.B., Gibson C.J., Lin A.E., Taub M.A., Aguet F., Ardlie K., Mitchell B.D., Barnes K.C., Moscati A., Fornage M., Redline S., Psaty B.M., Silverman E.K., Weiss S.T., Palmer N.D., Vasan R.S., Burchard E.G., Kardia S.L., He J., Kaplan R.C., Smith N.L., Arnett D.K., Schwartz D.A., Correa A., de Andrade M., Guo X., Konkle B.A., Custer B., Peralta J.M., Gui H., Meyers D.A., McGarvey S.T., Chen I.Y., Shoemaker M.B., Peyser P.A., Broome J.G., Gogarten S.M., Wang F.F., Wong Q., Montasser M.E., Daya M., Kenny E.E., North K.E., Launer L.J., Cade B.E., Bis J.C., Cho M.H., Lasky-Su J., Bowden D.W., Cupples L.A., Mak A.C., Becker L.C., Smith J.A., Kelly T.N., Aslibekyan S., Heckbert S.R., Tiwari H.K., Yang I.V., Heit J.A., Lubitz S., Johnsen J.M., Curran J.E., Wenzel S.E., Weeks D.E., Rao D.C., Darbar D., Moon J.Y., Tracy R.P., Buth E.J., Rafaels N., Loos R.J., Durda P., Liu Y., Hou L., Lee J., Kachroo P., Freedman B.I., Levy D., Bielak L.F., Hixson J.E., Floyd J.S., Whitsel E.A., Ellinor P.T., Irvin M.R., Fingerlin T.E., Raffield L.M., Armasu S.M., Wheeler M.M., Sabino E.C., Blangero J., Williams L.K., Levy B.D., Huey-Herng Sheu W., Roden D.M., Boerwinkle E., Manson J.E., Mathias R.A., Desai P., Taylor K.D., Johnson A.D., Auer P.L., Kooperberg C., Laurie C.C., Blackwell T.W., Smith A.V., Zhao H., Lange E., Lange L., Rich S.S., Rotter J.I., Wilson J.G., Scheet P., Kitzman J.O., Lander E.S., Engreitz J.M., Ebert B.L., Reiner A.P., Jaiswal S., Abecasis G., Sankaran V.G., Kathiresan S, & Natarajan P. (2020). Inherited Causes of Clonal Hematopoiesis in 97,691 TOPMed Whole Genomes. Nature, 586(7831), 763-768.
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6

Clonogenic Assay of CD34+ HSPCs

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3 days RNP post-electroporation, 500 CD34+ HSPCs were plated in 1ml methylcellulose media (#H4034, Stem Cell Technologies). Primary CFU-C colonies were counted after 14 days. For the colony replating experiments, 2 weeks after the primary plating, the colonies from two plates were pooled, washed with PBS, and the cells were plated in new methylcellulose media at 25000 cells/ml for an additional 14 days. Images of primary and secondary colonies were taken using StemVision (Stem Cell technologies) using manufacturer’s recommendations.
Bao E.L., Nandakumar S.K., Liao X., Bick A.G., Karjalainen J., Tabaka M., Gan O.I., Havulinna A., Kiiskinen T., Lareau C.A., de Lapuente Portilla A.L., Li B., Emdin C., Codd V., Nelson C.P., Walker C.J., Churchhouse C., de la Chapelle A., Klein D.E., Nilsson B., Wilson P.W., Cho K., Pyarajan S., Gaziano J.M., Samani N.J., Regev A., Palotie A., Neale B.M., Dick J.E., Natarajan P., O’Donnell C.J., Daly M.J., Milyavsky M., Kathiresan S, & Sankaran V.G. (2020). Inherited myeloproliferative neoplasm risk impacts hematopoietic stem cells. Nature, 586(7831), 769-775.
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7

Colony-Forming Cell Assay Protocol

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The colony-forming cell (CFC) generation assay was performed as previously described68 (link). Briefly, sorted cells were cultured on irradiated OP9DLL1 (a kind gift from Juan-Carlos Zúñiga-Pflücker) monolayers in Alpha MEM (ThermoFisher, 12000063) supplemented with 20% FBS (HyClone), 1% penicillin–streptomycin 2 mM l-glutamine, 30 ng ml−1 thrombopoietin (TPO) (Miltenyi Biotec, 130-095-747), 10 ng ml−1 BMP4, 50 ng ml−1, 25 ng ml−1 IGF1, 10 ng ml−1 IL11, 10 ng ml−1 FLT3L and 4 U ml−1 EPO. After 5 days, cells were collected using 0.25% trypsin–EDTA (Thermo Fisher Scientific, 25-200-056) for 3 min at 37 °C. Cells were then filtered through a 40 μM filter and seeded on methylcellulose medium (STEMCELL Technologies, H4034). Cells were seeded on methylcellulose supplemented with 150 µg ml−1 transferrin, 50 ng ml−1 TPO, 10 ng ml−1 VEGF, 10 ng ml−1 IL6, 50 ng ml−1 IGF1, 5 ng ml−1 IL11 and 4 U ml−1 EPO. Colonies’ number and morphology were evaluated after 15 days by light microscopy.
Scarfò R., Randolph L.N., Abou Alezz M., El Khoury M., Gersch A., Li Z.Y., Luff S.A., Tavosanis A., Ferrari Ramondo G., Valsoni S., Cascione S., Didelon E., Passerini L., Amodio G., Brandas C., Villa A., Gregori S., Merelli I., Freund J.N., Sturgeon C.M., Tavian M, & Ditadi A. (2024). CD32 captures committed haemogenic endothelial cells during human embryonic development. Nature Cell Biology, 26(5), 719-730.
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8

Isolation and Differentiation of Erythroid Cells

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Mononuclear cells were isolated from cord blood (CRB Saint- Louis Hospital, Paris, France) or adult bone marrow donors with written informed consent for research use, in accordance with the Declaration of Helsinki. The ethics evaluation committee of INSERM, the Institutional Review Board (IRB00003888), approved our research project (n. 16-319). Mononucleated cells were separated by Ficoll-Paque (Life Technologies) and purified with a human CD34 MicroBead Kit (cat. n. 130100453, Miltenyi Biotec). A two-step culture method for cell expansion and erythroid differentiation was used to obtain highly stage-enriched erythroblast populations.36 (link),37 (link) Briefly, CD34+ cells (purity 94-98%) were cultured for 7 days with interleukin (IL)6, IL3 (10 ng/mL), and stem cell factor (SCF, 100 ng/mL) to expand hematopoietic progenitors.37 (link) CD36+ erythroid progenitors were purified using magnetic microbeads (CD36 FA6.152 from Beckman Coulter and anti-mouse IgG1 MicroBeads from Miltenyi Biotec). Then, erythropoietin (EPO) was added for 10-18 days to allow erythroid differentiation. For assays of colony-forming cells (CFC, counted at 11-14 days), CD34+ hematopoietic stem cells were seeded in methylcellulose (H4034, StemCell Technologies). TGFbRI inhibitor SB431542 was from Selleckchem (cat. n. S1067)38 (link) and TGFb was from Peprotech (cat. n. 100-21).
Astori A., Matherat G., Munoz I., Gautier E.F., Surdez D., Zermati Y., Verdier F., Zaidi S., Feuillet V., Kadi A., Lauret E., Delattre O., Lefèvre C., Fontenay M., Ségal-Bendirdjian E., Dusanter-Fourt I., Bouscary D., Hermine O., Mayeux P, & Pendino F. (2020). The epigenetic regulator RINF (CXXC5) maintains SMAD7 expression in human immature erythroid cells and sustains red blood cell expansion. Haematologica, 107(1), 268-283.
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9

Quantifying Hematopoietic Progenitor Cells

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The CFU-C assays were performed using complete methylcellulose-based medium (MethoCult GF M3434 or H4034; Stem Cell Technologies) according to the manufacturer’s instructions. The colony numbers and morphology were assessed after 14 d of culture. All cultures were incubated at 37°C in a 5% CO2 humidified atmosphere.
Liu M., Wu C., Luo S., Hua Q., Chen H.T., Weng Y., Xu J., Lin H., Wang L., Li J., Zhu L., Guo Z., Zhuang S.M., Kang T, & Zheng L. (2022). PERK reprograms hematopoietic progenitor cells to direct tumor-promoting myelopoiesis in the spleen. The Journal of Experimental Medicine, 219(4), e20211498.
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10

HSPC Colony Forming Assay

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Three days after RNP electroporation, 500 CD34+ HSPCs were plated in 1 ml methylcellulose medium (H4034, Stem Cell Technologies) in triplicate unless otherwise noted. Primary colonies were counted after 14 d.
Voit R.A., Tao L., Yu F., Cato L.D., Cohen B., Fleming T.J., Antoszewski M., Liao X., Fiorini C., Nandakumar S.K., Wahlster L., Teichert K., Regev A, & Sankaran V.G. (2022). A genetic disorder reveals a hematopoietic stem cell regulatory network co-opted in leukemia. Nature Immunology, 24(1), 69-83.
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