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Cpg b odn2006

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CpG-B ODN2006 is an oligonucleotide that contains unmethylated cytosine-guanine (CpG) motifs. It functions as a potent activator of the innate immune system by binding to Toll-like receptor 9 (TLR9).

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Lab products found in correlation

Sodium pyruvate, by Thermo Fisher Scientific (3 mentions) L glutammine, by Thermo Fisher Scientific (2 mentions) Rpmi 1640, by Euroclone (2 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (2 mentions) Rpmi 1640, by Thermo Fisher Scientific (1 mentions) Phosphate buffered saline (pbs), by Euroclone (1 mentions) Penicillin streptomicin 100x, by Euroclone (1 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions) Penicillin streptomycin 100, by Euroclone (1 mentions) L glutamine, by Thermo Fisher Scientific (1 mentions) Hyclone fbs, by Cytiva (1 mentions)

4 protocols using cpg b odn2006

1

CpG-B ODN2006 Stimulation Assay

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Cells were cultured at a concentration of 2.5 × 106 cells/mL in 96-multiwell plates (Becton Dickinson, San Jose, CA, USA) and cultured for different time points as described in figure legends. CpG-B ODN2006 (Hycult Biotech) was used at 0.35 μM concentration. Complete medium was prepared as follows: RPMI-1640 (Gibco BRL, Life Technologies), 10% FBS, 1% L-Glutammine (Gibco BRL); 1% Antibiotics/Antimicotics (Gibco BRL), 1% sodium pyruvate (Gibco BRL).
Farroni C., Marasco E., Marcellini V., Giorda E., Valentini D., Petrini S., D'Oria V., Pezzullo M., Cascioli S., Scarsella M., Ugazio A.G., De Vincentiis G.C., Grimsholm O, & Carsetti R. (2018). Dysregulated miR-155 and miR-125b Are Related to Impaired B-cell Responses in Down Syndrome. Frontiers in Immunology, 9, 2683.
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2

CFSE-Based Proliferation Assay for PBMCs

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PBMCs were loaded with Carboxyfluorescein succinimidyl ester (CFSE, Life Technologies). Briefly, 1x106 cells/ml were resuspended in PBS (Euroclone) with 1% FBS (Gibco BRL) and loaded with 1μM CFSE for 20 min at 37°C. After washing, cells (at a concentration of 5x106 cells/ml) were stimulated with 0.35 μM of TLR9 agonist CpG-B ODN2006 (Hycult Biotech) in complete medium for 5 days at 37°C. Complete medium was prepared as follows: RPMI-1640 (Euroclone), 10% heat inactivated fetal bovine serum (FBS, Hyclone Laboratories), 1% L-Glutammine (Gibco BRL); 1% Penicillin/Streptomicin 100X (Euroclone), 1% sodium pyruvate (Gibco BRL).
Ciocca M., Zaffina S., Fernandez Salinas A., Bocci C., Palomba P., Conti M.G., Terreri S., Frisullo G., Giorda E., Scarsella M., Brugaletta R., Vinci M.R., Magnavita N., Carsetti R, & Piano Mortari E. (2021). Evolution of Human Memory B Cells From Childhood to Old Age. Frontiers in Immunology, 12, 690534.
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3

Measuring B-cell Proliferation via CFSE

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B-cell proliferation was determined according to ref. (41 (link)). Briefly, 1×106 cells/ml splenocytes from pristane-treated mice were loaded with 1μM CFSE (Life Technologies) in PBS/1% FCS for 20min at 37°C and were blocked by washing with IMDM/20%FCS. 2x106 cells/ml were then cultured in IMDM/20%FCS under stimulation with 0.35μM CpG-B ODN2006 (Hycult Biotech) for 5 days. B-cell proliferation was evaluated by tracking dilution of CFSE within the B220+CD19+ flow-gate.
Czeh M., Stäble S., Krämer S., Tepe L., Talyan S., Carrelha J., Meng Y., Heitplatz B., Schwabenland M., Milsom M.D., Plass C., Prinz M., Schlesner M., Andrade-Navarro M.A., Nerlov C., Jacobsen S.E., Lipka D.B, & Rosenbauer F. (2021). DNMT1 deficiency impacts on plasmacytoid dendritic cells in homeostasis and autoimmune disease. Journal of immunology (Baltimore, Md. : 1950), 208(2), 358-370.
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4

Polyclonal Stimulation of PBMCs

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As we published before [14 (link)], in order to induce antibody production from MBCs, we polyclonally stimulated PBMCs with CpG. Briefly, PBMCs were cultured in complete medium at a concentration of 1 × 106 cells/mL. Complete medium was prepared as follows: RPMI-1640 (Euroclone, Milan, Italy), 10% heat inactivated HyClone FBS (Hyclone Laboratories, Logan, UT, USA), 1mM L-Glutamine (GIBCO BRL, New York, NY, USA); 1× Penicillin/Streptomycin 100× (Euroclone, Milan, Italy), 1% sodium pyruvate (GIBCO BRL, New York, NY, USA). Cells were stimulated for 5 days with 0.35 µM TLR9 agonist CpG-B ODN2006 (Hycult Biotech, Uden, The Netherland) plus 20 ng/mL rhIL-21 (Milteny, Bergisch Gladbach, Germany) and 20 ng/mL rhIL-4 (Milteny, Bergisch Gladbach, Germany).
Piano Mortari E., Russo C., Vinci M.R., Terreri S., Fernandez Salinas A., Piccioni L., Alteri C., Colagrossi L., Coltella L., Ranno S., Linardos G., Agosta M., Albano C., Agrati C., Castilletti C., Meschi S., Romania P., Roscilli G., Pavoni E., Camisa V., Santoro A., Brugaletta R., Magnavita N., Ruggiero A., Cotugno N., Amodio D., Ciofi Degli Atti M.L., Giorgio D., Russo N., Salvatori G., Corsetti T., Locatelli F., Perno C.F., Zaffina S, & Carsetti R. (2021). Highly Specific Memory B Cells Generation after the 2nd Dose of BNT162b2 Vaccine Compensate for the Decline of Serum Antibodies and Absence of Mucosal IgA. Cells, 10(10), 2541.
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