Ril 2

Manufactured by BD

Sourced in United States

The RIL-2 is a laboratory equipment designed for the rapid and efficient isolation of ribonucleic acid (RNA) from various biological samples. It utilizes a proprietary extraction method to ensure high-quality RNA yields.

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Lab products found in correlation

7 protocols using ril 2

SARS-CoV-2 Spike Protein Peptide Stimulation

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PBMCs were isolated from heparinized whole blood using Ficoll–Paque (GE Healthcare, Singapore). Then, PBMCs from I-I-I were treated with the peptide pool containing 384 15-mer peptides spanning the antigen region of spike (S) protein (250 nM per peptide) in presence of 10 U/mL recombinant interleukin-2 (rIL-2) and 1 μM GolgiPlug (BD Biosciences, San Diego, CA) for 16 h at 37 °C, 5% CO₂. PBMCs from BA.5 infection were treated with the peptide pool containing 487 15-mer peptides spanning the antigen region of spike (S), membrane (M), nucleocapsid (N), and envelope (E) proteins (250 nM per peptide) in presence of 10 U/mL rIL-2 and 1 μM GolgiPlug (BD Biosciences, San Diego, CA) for 16 h at 37 °C, 5% CO₂. RPMI 1640 medium (Gibco, Waltham, MA) supplemented with 10% heat-inactivated FBS (Biological Industries, Israel Beit-Haemek), 100 U/mL penicillin (Gibco, Waltham, MA), 0.1 mg/mL streptomycin (Gibco, Waltham, MA), 10 U/mL rIL-2, and 0.01% DMSO (Sigma, Saint Louis, MO) was used as subtraction control.

Cui T., Su X., Sun J., Liu S., Huang M., Li W., Luo C., Cheng L., Wei R., Song T., Sun X., Luo Q., Li J., Su J., Deng S., Zhao J., Zhao Z., Zhong N, & Wang Z. (2023). Dynamic immune landscape in vaccinated-BA.5-XBB.1.9.1 reinfections revealed a 5-month protection-duration against XBB infection and a shift in immune imprinting. eBioMedicine, 99, 104903.

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PBMCs Stimulation for Immune Analysis

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PBMCs were isolated from heparinized whole blood by density-gradient sedimentation using Ficoll-Paque according to the manufacturer’s instructions (GE Healthcare, 17-1440-02). PBMCs (5 × 10⁵) were cultured in RPMI 1640 medium (Gibco, Waltham, MA, USA) supplemented with 10% heat-inactivated FBS (Biological Industries, Israel Beit-Haemek), 100 U/mL penicillin (Gibco, Waltham, MA, USA), and 0.1 mg/mL streptomycin (Gibco, Waltham, MA, USA). PBMCs were treated overnight using a peptide pool containing 56 15-mer peptides (2 μM per peptide) in the presence of 10 U/mL rIL-2 and 1 µM GolgiPlug (BD Biosciences, San Diego, CA, USA) at 37 °C and 5% CO₂.

Zhao Z., Cui T., Huang M., Liu S., Su X., Li G., Song T., Li W., Zhong N., Xu M., Yang X., Huang W, & Wang Z. (2022). Heterologous boosting with third dose of coronavirus disease recombinant subunit vaccine increases neutralizing antibodies and T cell immunity against different severe acute respiratory syndrome coronavirus 2 variants. Emerging Microbes & Infections, 11(1), 829-840.

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Peptide Stimulation of PBMC Cultures

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Peripheral blood mononuclear cells (PBMCs) were isolated from heparinized whole blood by density-gradient sedimentation using Ficoll-Paque according to the manufacturer’s instructions (GE Healthcare, 17-1440-03). 5×10⁵ PBMCs were cultured in RPMI 1640 medium (Gibco) supplemented with 10% heat inactivated FBS (Biological Industries, Israel Beit-Haemek), 100 U/mL penicillin (Gibco) and 0.1 mg/mL streptomycin (Gibco). The PBMCs were treated with the peptide pool containing 383 15-mer peptides at 250 nM/each peptide in the presence of 20 U/mL rIL-2 and 1 μM GolgiPlug (BD Biosciences, San Diego, CA) overnight at 37°C, 5% CO₂ (Wang et al., 2021 (link)). The approach of using a large peptide pool to stimulate PBMCs was based on that developed by Chevalier et al. and was previously validated for CMV peptides (Chevalier et al., 2015 (link)).

Zhang J., Cao J., Zheng R., Yu M., Lin Z., Wang C., McCluskey J., Yang J., Chen Z., Corbett A.J., Cao P., Mo W, & Wang Z. (2022). The establishment of a cytomegalovirus -specific CD8+ T-cell threshold by kinetic modeling for the prediction of post-hemopoietic stem cell transplant reactivation. iScience, 25(11), 105340.

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Peptide-Stimulated PBMC Activation Assay

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PBMCs were isolated from heparinized whole blood by density gradient sedimentation using Ficoll-Paque according to the manufacturer’s instructions (GE Healthcare, 17-1440-02). The PBMCs (5 × 10⁵) were then cultured in complete RPMI (c-RPMI, RPMI 1640 medium (Gibco) enriched with supplements, including 10% heat-inactivated FBS (Biological Industries, Israel Beit-Haemek), 100 μM MEM nonessential amino acids (Gibco), 100 U/mL penicillin (Gibco), 0.1 mg/mL streptomycin (Gibco), 2 mM L-glutamine (Gibco), 25 mM HEPES (Gibco), 55 μM 2-mercaptoethanol (Gibco) and 1 mM sodium pyruvate (Gibco)). The PBMCs were treated with the peptide pool containing 487 15-mer peptides (250 nM of each peptide) in the presence of 10 U/mL rIL-2 and 1 μM GolgiPlug (BD Biosciences, San Diego, CA, USA) for 16 h at 37 °C with 5% CO₂. The approach of using a large peptide pool to stimulate PBMCs was based on that developed by Chevalier et al.^{34 (link)} and was validated.

Wang J., Li K., Mei X., Cao J., Zhong J., Huang P., Luo Q., Li G., Wei R., Zhong N., Zhao Z, & Wang Z. (2022). SARS-CoV-2 vaccination-infection pattern imprints and diversifies T cell differentiation and neutralizing response against Omicron subvariants. Cell Discovery, 8, 136.

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Isolation and Stimulation of PBMCs

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PBMCs were isolated from heparinized whole blood by density-gradient sedimentation using Ficoll-Paque according to the manufacturer’s instructions (GE Healthcare, 17-1440-02). 1 × 10⁶ PBMCs were cultured in RPMI 1640 medium (Gibco) supplemented with 10% heat-inactivated FBS (Biological Industries, Israel Beit-Haemek), 100 U/ml penicillin (Gibco) and 0.1 mg/ml streptomycin (Gibco). The PBMCs were treated with the peptide pool containing 447 15-mer peptides and 110 18-mer peptides at 125 nM/each peptide in the presence of 10 U/ml rIL-2 and 1 µM GolgiPlug (BD Biosciences, San Diego, CA) overnight at 37 °C, 5% CO₂. The approach of using a large peptide pool to stimulate PBMCs was based on that developed by Chevalier M. F. et al.^{29 (link)} and was validated for CMV peptides.

Wang Z., Yang X., Zhong J., Zhou Y., Tang Z., Zhou H., He J., Mei X., Tang Y., Lin B., Chen Z., McCluskey J., Yang J., Corbett A.J, & Ran P. (2021). Exposure to SARS-CoV-2 generates T-cell memory in the absence of a detectable viral infection. Nature Communications, 12, 1724.

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Murine Treg Cell Differentiation

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Naive T cells (CD4⁺CD62L⁺) from spleen and lymph nodes of female C57BL/6 mice, purified using magnetic beads (Mylteni Biotec, Bergisch Gladbach, NRW, Germany), were distributed in plates previously coated overnight with monoclonal antibody anti-CD3 (5 μg/mL, BD Pharmingen, San Diego, CA, USA) in the presence of anti-CD28 (1 μg/mL), rTGF-β (3 ng/mL, BD Pharmingen, San Diego, CA, USA) and rIL-2 (10 ng/mL, BD Pharmingen, San Diego, CA, USA), treated or not with ArtC (0.01, 0.1, 5, 10, 20, 50 or 100 μM). For Treg cell differentiation, cells were cultured for 96 h, at 37 °C and 5% CO₂.

Martins N.S., de Campos Fraga-Silva T.F., Correa G.F., Boko M.M., Ramalho L.N., Rodrigues D.M., Hori J.I., Costa D.L., Bastos J.K, & Bonato V.L. (2021). Artepillin C Reduces Allergic Airway Inflammation by Induction of Monocytic Myeloid-Derived Suppressor Cells. Pharmaceutics, 13(11), 1763.

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Cytolytic Activity Assay of CTLs

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Splenocytes were cultured with WT1_126-134 peptide -coated DCs at a 20:1 ratio for 24 h after which, cells were split and cultured in medium supplemented with murine rIL-2 (0.3 ng/ml) (BD Biosciences). The cytolytic activity of CTLs against ID8-T tumor cells was analyzed 5 days later by a standard 4-h ⁵¹Cr-release assay. The percentage of specific lysis was calculated as follows: ([cpm experimental release - cpm spontaneous release]/[cpm maximum release cpm spontaneous release]) × 100. Maximum release was determined from the supernatants of cells that were lysed by the addition of 5% Triton X-100. Spontaneous release was determined from target cells incubated with medium only.

Gil M., Komorowski M.P., Seshadri M., Rokita H., McGray A.J., Opyrchal M., Odunsi K.O, & Kozbor D. (2014). CXCL12/CXCR4 Blockade by Oncolytic Virotherapy Inhibits Ovarian Cancer Growth by Decreasing Immunosuppression and Targeting Cancer Initiating Cells. Journal of immunology (Baltimore, Md. : 1950), 193(10), 5327-5337.

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