RosetteSep® Human B Cell Enrichment Cocktail

Procedures and instruction manuals:

•  PRODUCT INFORMATION SHEET - 15064, Lot# All:28573-PIS_1_1_0.pdf
•  PRODUCT INFORMATION SHEET - 15024, Lot# All:28573-PIS_1_1_0.pdf

Educational resources:

•  VIDEO: RosetteSep™: Cell Isolation Directly From Whole Blood Without Columns or Magnets
•  VIDEO: RosetteSep®: A Guide to Isolating NK Cells Directly from Whole Blood
•  BROCHURE: One-Step Cell Enrichment Directly From Whole Blood
•  BROCHURE: Cell Separation for Autoimmunity Research
•  WALLCHART: Antigen Processing and Presentation Wallchart
•  BROCHURE: Ready Sep Go – Cell Separation In As Little As 25 Minutes
•  SCIENTIFIC POSTER: One-step enrichment of leukocyte subsets directly in the blood collection tube
•  VIDEO: RosetteSep® for HLA Analysis
•  BROCHURE: Fast and Easy Isolation of B Cells

MSDS:

•  15014C_15024C-MSDS.pdf

FAQS:

• 
  HOW ROSETTESEP™ WORKS
  
  
  • Q. WHAT IS ROSETTESEP™?
    A. RosetteSep™ is a rapid cell separation procedure for the isolation of purified cells directly from whole blood, without columns or magnets.
  • Q. HOW DOES ROSETTESEP™ WORK?
    A. The antibody cocktail crosslinks unwanted cells to red blood cells (RBCs), forming rosettes. The unwanted cells then pellet with the free RBCs when centrifuged over a density centrifugation medium (e.g. Ficoll-Paque™ PLUS, Lymphoprep™).
  • Q. WHAT FACTORS AFFECT CELL RECOVERY?
    A. The temperature of the reagents can affect cell recovery. All reagents should be at room temperature (sample, density centrifugation medium, PBS, centrifuge) before performing the isolations. Layering can also affect recovery so be sure to carefully layer the sample to avoid mixing with the density centrifugation medium as much as possible. Be sure to collect the entire enriched culture without disturbing the RBC pellet. A small amount of density centrifugation medium can be collected without worry.
  
  CELL TYPES AND SAMPLE SOURCES
  
  
  • Q. WHICH CELL SAMPLES CAN ROSETTESEP™ BE USED WITH?
    A. RosetteSep™ can be used with leukapheresis samples, bone marrow or buffy coat, as long as: the concentration of cells does not exceed 5 x 10⁷ per mL (can dilute if necessary); and there are at least 20 RBCs for every nucleated cell (RBCs can be added if necessary).
  • Q. CAN ROSETTESEP™ BE USED WITH PREVIOUSLY FROZEN OR CULTURED CELLS?
    A. Yes. Cells should be re-suspended at 2 - 5 x 10⁷ cells / mL in PBS + 2% FBS. Fresh whole blood should be added at 250 µL per mL of sample, as a source of red cells.
  • Q. CAN ROSETTESEP™ BE USED TO ENRICH PROGENITORS FROM CORD BLOOD?
    A. Yes. Sometimes cord blood contains immature nucleated red cells that have a lower density than mature RBCs. These immature red cells do not pellet over Ficoll™, which can lead to a higher RBC contamination than peripheral blood separations.
  • Q. DOES ROSETTESEP™ WORK WITH MOUSE CELLS?
    A. No, but we have developed EasySep™, a magnetic-based cell isolation system which works with mouse and other non-human species.
  
  ANTI-COAGULANTS
  
  
  • Q. WHICH ANTICOAGULANT SHOULD BE USED WITH ROSETTESEP™?
    A. Peripheral blood should be collected in heparinized Vacutainers. Cord blood should be collected in ACD.
  • Q. SHOULD THE ANTICOAGULANT BE WASHED OFF BEFORE USING ROSETTESEP™?
    A. No, the antibody cocktail can be added directly to the sample.

This product has been used in:

1. Amy E Gilbert et al. Monitoring the systemic human memory B cell compartment of melanoma patients for anti-tumor IgG antibodies.PLoS One 6 (4) e19330 (2011)
2. Anita Parmigiani et al. Interleukin-21 and cellular activation concurrently induce potent cytotoxic function and promote antiviral activity in human CD8 T cells.Hum Immunol (October 25, 2010)
3. Lee A Honigberg et al. The Bruton tyrosine kinase inhibitor PCI-32765 blocks B-cell activation and is efficacious in models of autoimmune disease and B-cell malignancy.Proc Natl Acad Sci U S A 107 (29) 13075-13080 (July 20, 2010)
4. Thibaut Brugat et al. Telomere dysfunction-induced foci arise with the onset of telomeric deletions and complex chromosomal aberrations in resistant chronic lymphocytic leukemia cells.Blood 116 (2) 239-249 (July 15, 2010)
5. Isabelle Isnardi et al. Complement receptor 2/CD21- human naive B cells contain mostly autoreactive unresponsive clones.Blood 115 (24) 5026-5036 (June 17, 2010)
6. Michela Frenquelli et al. MicroRNA and proliferation control in chronic lymphocytic leukemia: functional relationship between miR-221/222 cluster and p27.Blood 115 (19) 3949-3959 (May 13, 2010)
7. Zibo Song et al. Activities of SYK and PLCgamma2 predict apoptotic response of CLL cells to SRC tyrosine kinase inhibitor dasatinib.Clin Cancer Res 16 (2) 587-599 (January 15, 2010)
8. Jonathan Zuccolo et al. Efficient isolation of highly purified tonsil B lymphocytes using RosetteSep with allogeneic human red blood cells.BMC Immunol 10 30 (2009)
9. Jennifer L Costantini et al. TAPP2 links phosphoinositide 3-kinase signaling to B-cell adhesion through interaction with the cytoskeletal protein utrophin: expression of a novel cell adhesion-promoting complex in B-cell leukemia.Blood 114 (21) 4703-4712 (November 19, 2009)
10. Marco Herling et al. High TCL1 levels are a marker of B-cell receptor pathway responsiveness and adverse outcome in chronic lymphocytic leukemia.Blood 114 (21) 4675-4686 (November 19, 2009)
11. David M Lucas et al. The novel plant-derived agent silvestrol has B-cell selective activity in chronic lymphocytic leukemia and acute lymphoblastic leukemia in vitro and in vivo.Blood 113 (19) 4656-4666 (May 7, 2009)
12. Rosa Lapalombella et al. The humanized CD40 antibody SGN-40 demonstrates pre-clinical activity that is enhanced by lenalidomide in chronic lymphocytic leukaemia.Br J Haematol 144 (6) 848-855 (March 2009)
13. Hai-Jun Zhou et al. Nuclear CD40 interacts with c-Rel and enhances proliferation in aggressive B-cell lymphoma.Blood 110 (6) 2121-2127 (September 15, 2007)
14. Sheng Chen et al. Modulatory effects of 1,25-dihydroxyvitamin D3 on human B cell differentiation.J Immunol 179 (3) 1634-1647 (August 1, 2007)
15. Maxime Herv?? et al. CD40 ligand and MHC class II expression are essential for human peripheral B cell tolerance.J Exp Med 204 (7) 1583-1593 (July 9, 2007)
16. Montserrat Barragan et al. Regulation of Akt/PKB by phosphatidylinositol 3-kinase-dependent and -independent pathways in B-cell chronic lymphocytic leukemia cells: role of protein kinase C{beta}J Leukoc Biol 80 (6) 1473-1479 (December 1, 2006)
17. Rengasamy Asokan et al. Characterization of human complement receptor type 2 (CR2/CD21) as a receptor for IFN-alpha: a potential role in systemic lupus erythematosus.J Immunol 177 (1) 383-394 (July 1, 2006)
18. Diana Starlets et al. Cell-surface CD74 initiates a signaling cascade leading to cell proliferation and survival.Blood 107 (12) 4807-4816 (June 15, 2006)
19. Lingchen Fu et al. Constitutive NF-kappaB and NFAT activation leads to stimulation of the BLyS survival pathway in aggressive B-cell lymphomas.Blood 107 (11) 4540-4548 (June 1, 2006)
20. Tara M C Hornell et al. Human dendritic cell expression of HLA-DO is subset specific and regulated by maturation.J Immunol 176 (6) 3536-3547 (March 15, 2006)
21. Sandra Weller et al. Human blood IgM "memory" B cells are circulating splenic marginal zone B cells harboring a prediversified immunoglobulin repertoire.Blood 104 (12) 3647-3654 (December 1, 2004)
22. Yen-Shing Ng et al. Bruton's tyrosine kinase is essential for human B cell tolerance.J Exp Med 200 (7) 927-934 (October 4, 2004)
23. Heike Wulff et al. K+ channel expression during B cell differentiation: implications for immunomodulation and autoimmunity.J Immunol 173 (2) 776-786 (July 15, 2004)
24. Guo-Qiang Chen et al. Methylated metabolites of arsenic trioxide are more potent than arsenic trioxide as apoptotic but not differentiation inducers in leukemia and lymphoma cells.Cancer Res 63 (8) 1853-1859 (April 15, 2003)
25. Anna Zaninoni et al. Cytokine modulation of nuclear factor-kappaB activity in B-chronic lymphocytic leukemia.Exp Hematol 31 (3) 185-190 (March 2003)
26. Melissa L Precopio et al. Differential kinetics and specificity of EBV-specific CD4+ and CD8+ T cells during primary infection.J Immunol 170 (5) 2590-2598 (March 1, 2003)
27. Lydie B??niguel et al. Differential production in vitro of antigen specific IgG1, IgG3 and IgA: a study in Schistosoma haematobium infected individuals.Parasite Immunol 25 (1) 39-44 (January 2003)
28. Claire E Hirst et al. The intracellular granzyme B inhibitor, proteinase inhibitor 9, is up-regulated during accessory cell maturation and effector cell degranulation, and its overexpression enhances CTL potency.J Immunol 170 (2) 805-815 (January 15, 2003)

FACS Histogram Results With RosetteSep® Human B Cell Enrichment Cocktail

RosetteSep® Procedure