Rosettesep cd4 t cell enrichment cocktail

Manufactured by STEMCELL

Sourced in Canada

RosetteSep CD4+ T cell enrichment cocktail is a cell separation product designed to isolate CD4+ T cells from whole blood or other cell suspensions. It contains antibodies that crosslink unwanted cells to red blood cells, facilitating the removal of non-target cells during centrifugation.

Automatically generated - may contain errors

Lab products found in correlation

Cd25 microbeads, by Miltenyi Biotec (2 mentions) Rneasy mini kit, by Qiagen (1 mentions) Ficoll plaque plus, by GE Healthcare (1 mentions) Genechip human transcriptome array 2.0 microarrays, by Thermo Fisher Scientific (1 mentions) Ncounter xt assay, by NanoString (1 mentions) Ficoll paque, by Cytiva (1 mentions) Facsaria, by BD (1 mentions) Penicillin, by Thermo Fisher Scientific (1 mentions) Rna clean and concentrator kit, by Zymo Research (1 mentions) Glutamax, by Thermo Fisher Scientific (1 mentions) Neon transfection system, by Thermo Fisher Scientific (1 mentions) Engen sgrna synthesis kit, by New England Biolabs (1 mentions) Immunocult, by STEMCELL (1 mentions) Facsaria flow cytometer, by BD (1 mentions)

Close spelling variants of this product

RosetteSep (157 citations) RosetteSep CD4 Enrichment Cocktail (2 citations) RosetteSep Human CD4+ T Cells Enrichment Cocktail (2 citations)

7 protocols using rosettesep cd4 t cell enrichment cocktail

Isolation and Characterization of Primary CD4+ T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

For subsequent total RNA-Seq analysis, cryopreserved primary CD4⁺ T cells that were viably frozen were obtained from four different healthy donors (AllCells, Inc., Emeryville, CA, USA) and thawed in RPMI with 20% human serum. Dead cells resulting from thawing frozen cells were removed using Viahance magnetic negative selection (Biophysics Assay Laboratory Inc., Worcester, MA, USA). For dose–response analysis, peripheral blood was isolated from two additional healthy donors by venipuncture according to the protocols approved by an institutional review board into polypropylene syringes containing sodium heparin. Primary CD4⁺ T cells were isolated using the RosetteSep CD4⁺ T cell enrichment cocktail (StemCell Technologies Inc., Vancouver, Canada). Aliquots taken from CD4⁺ T cell samples were subjected to flow cytometry to assess purity (i.e., >95% cells expressing CD4).

White C.H., Beliakova-Bethell N., Lada S.M., Breen M.S., Hurst T.P., Spina C.A., Richman D.D., Frater J., Magiorkinis G, & Woelk C.H. (2018). Transcriptional Modulation of Human Endogenous Retroviruses in Primary CD4+ T Cells Following Vorinostat Treatment. Frontiers in Immunology, 9, 603.

+ Open protocol

+ Expand

CD4+ T Cell Transcriptome Analysis in SLE

Check if the same lab product or an alternative is used in the 5 most similar protocols

CD4⁺ T cells were isolated by negative selection from whole blood using the RosetteSep CD4⁺ T cell enrichment cocktail (Stemcell Technologies) and Ficoll-Plaque Plus (GE Healthcare) by gravity centrifugation. RNA isolated from eight HC and seven SLE patients (one individual with a pair of samples 6 months apart) using the RNeasy Mini Kit (Qiagen) was prepared for GeneChip^™ Human Transcriptome Array 2.0 microarrays (Affymetrix, Thermo Fisher Scientific) with fragmentation, labeling, hybridization and scanning as a single batch. Microarray CEL files were processed by Partek Genomics Suite 6.6 (Partek) using the RMA normalization procedure. Gene set enrichment analysis was carried out with GSEA software [8 (link)] on RMA normalized values using the curated KEGG database v6.0 or HALLMARKS v6.0. A custom gene expression panel (Supplemental Table 1) was designed for nCounter® XT Assay (NanoString Technologies). 50 ng of RNA from CD4⁺ T cell samples were hybridized to cartridges and scanned according to manufacturer instructions. Data was analyzed with nSolver 2.5 with geometric mean normalization to housekeeping genes as well as to positive and negative spike-in controls.

Perry D.J., Titov A.A., Sobel E.S., Brusko T.M, & Morel L. (2020). Immunophenotyping Reveals Distinct Subgroups of Lupus Patients Based on their Activated T Cell Subsets. Clinical immunology (Orlando, Fla.), 221, 108602.

+ Open protocol

+ Expand

Isolation and Characterization of Human Regulatory T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

CD4 positive T cells were isolated directly from fresh leukopacks (American Red Cross, Columbus, OH) by 30-minute incubation with Rosette Sep CD4 T cell enrichment cocktail (Stem Cell Technologies, Vancouver, BC), followed by Ficoll-Paque (Amersham Pharmacia Biotech, Uppsala, Sweden) density gradient centrifugation. CD4⁺CD25⁺ cells were then isolated by positive selection using CD25 microbeads (Miltenyi Biotec, Auburn, CA). Purity of isolated CD4⁺CD25⁺ cells and FoxP3 expression were evaluated by flow cytometry using the Regulatory T Cell Staining Kit (CD4 APC, CD25 FITC, and FoxP3 PE; eBioscience, San Diego, CA).

del Campo S.E., Levine K.M., Mundy-Bosse B.L., Grignol V.P., Fairchild E.T., Campbell A.R., Trikha P., Mace T.A., Paul B.K., Jaime-Ramirez A.C., Markowitz J., Kondadasula S.V., Guenterberg K.D., McClory S., Karpa V.I., Pan X., Olencki T.E., Monk J.P., Mortazavi A., Tridandapani S., Lesinski G.B., Byrd J.C., Caligiuri M.A., Shah M.H, & Carson WE I.I.I. (2015). The Raf Kinase Inhibitor Sorafenib Inhibits JAK-STAT Signal Transduction in Human Immune Cells. Journal of immunology (Baltimore, Md. : 1950), 195(5), 1995-2005.

+ Open protocol

+ Expand

Genetic Modification of Human Regulatory T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

CD4+CD25+ T cells were enriched from fresh human leukocyte cones by using a RosetteSep CD4+ T cell enrichment cocktail (Stemcell Technologies) and CD25 Microbeads (Miltenyi Biotech). CD4+CD25hiCD127lo Treg were then flow sorted using a FACSAria (BD Biosciences). Purified Treg were pre-activated in OpTmizer medium, supplemented with 1% penicillin and streptomycin, and 1% GlutaMAX (all Gibco) with 6.25 µl/ml CD3/CD28 tetramers (ImmunoCult; Stemcell Technologies) for 2 days with 1000 IU/ml IL-2.
sgRNAs were designed using http://crispr/mit/edu/ and synthesized using the EnGen sgRNA Synthesis Kit (NEB) and purified with the RNA Clean and Concentrator Kit (Zymo Research) according to the manufacturers’ instructions. Treg underwent electroporation using the Neon Transfection System (Invitrogen) (voltage: 2000 v, width 20 ms, 2 pulses) with 20 pmol Cas9 (Invitrogen) and 500 ng CTLA-4 specific sgRNA. Untreated control Treg did not receive sgRNA, Cas9 or undergo electroporation. Treg were expanded by co-plating with 4 × 10⁴ irradiated L cells as feeder cells, 100 ng/ml anti-CD3 (OKT3) and 1000 IU/ml IL-2 in 96-well plates. Treg were split as required and after at least 6 days Treg were rested for 24 h in fresh media with 100IU IL-2, then underwent quantification of total CTLA-4 levels by flow cytometry and used in stimulation assays, as above.

Halliday N., Williams C., Kennedy A., Waters E., Pesenacker A.M., Soskic B., Hinze C., Hou T.Z., Rowshanravan B., Janman D., Walker L.S, & Sansom D.M. (2020). CD86 Is a Selective CD28 Ligand Supporting FoxP3+ Regulatory T Cell Homeostasis in the Presence of High Levels of CTLA-4. Frontiers in Immunology, 11, 600000.

+ Open protocol

+ Expand

Isolation and Enrichment of CD4+ T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

PBMC were isolated by density gradient centrifugation as described in the Supplemental Methods. For the enrichment of CD4⁺ T cells, blood samples were mixed with RosetteSep CD4⁺ T cell enrichment cocktail (Stemcell Technologies) prior to centrifugation.

Diegelmann J, & Brand S. (2023). Identification of IL-27 as a novel regulator of major histocompatibility complex class I and class II expression, antigen presentation, and processing in intestinal epithelial cells. Frontiers in Immunology, 14, 1226809.

+ Open protocol

+ Expand

Isolation and Sorting of Naive CD4+ T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Before purification by cell sorting, CD4⁺ T cells were isolated from approximately 2 × 10⁹ elutriated lymphocytes by negative selection using RosetteSep CD4⁺ T cell enrichment cocktail (StemCell Technologies). Naïve CD4⁺ T cells from cord blood were sorted based on a phenotype of CD4⁺CD45RA⁺CD62L⁺CD25⁻CXCR3⁻CCR4⁻. In some experiments, cells were also purified based on staining with anti-CD161. For cells from adult blood, staining always included anti-CD4 plus anti-CD45RO. In addition, cells were stained with anti-CCR6 or anti-CXCR3 or anti-CCR4 or anti-CXCR5. All cell sorting was done using a FACS Aria flow cytometer (BD Biosciences). By post-sort analysis, the purity of populations was 95-99%.

Singh S.P., Zhang H.H., Tsang H., Gardina P.J., Myers T.G., Nagarajan V., Lee C.H, & Farber J.M. (2015). PLZF Regulates CCR6 and is Critical for the Acquisition and Maintenance of the Th17 Phenotype in Human Cells. Journal of immunology (Baltimore, Md. : 1950), 194(9), 4350-4361.

+ Open protocol

+ Expand

Isolation of Naïve CD4+ T Cells

Check if the same lab product or an alternative is used in the 5 most similar protocols

Leukocyte concentrates were obtained from healthy donors from the Centro Estatal de la Transfusión Sanguínea, Cuernavaca, Mexico. Peripheral blood mononuclear cells were obtained from these cellular concentrates through centrifugation with Ficoll-Hypaque gradient. Total CD4 + T cells were obtained using the RosetteSep CD4 + T cell enrichment cocktail (STEMCELL) and 1 ml of erythrocytes from the same donor. We depleted memory cells with an anti-CD45RO antibody (Tonbo) coupled to magnetic beads (Pierce) with the help of a magnetic rack. Naïve CD4 + T cells obtained this way were cultured in RPMI medium supplemented with 5% fetal bovine serum (FBS) at 37°C with 5% CO 2 . Cell preparations were routinely checked for purity and were at least 96% CD3 + CD4 + , CD45RO -, and 96% CCR7 + CD62L + .

Rodríguez-Jorge O., Kempis-Calanis L.A., Abou-Jaoudé W., Gutiérrez-Reyna D.Y., Hernandez C., Ramirez-Pliego O., Thomas-Chollier M., Spicuglia S., Santana M.A, & Thieffry D. (2019). Cooperation between T cell receptor and Toll-like receptor 5 signaling for CD4⁺ T cell activation. Science signaling, 12(577).

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!