Cd40l

Manufactured by Thermo Fisher Scientific

Sourced in United States, United Kingdom

CD40L is a recombinant human protein that functions as a ligand for the CD40 receptor. It is used in cell biology research to study immune system signaling pathways.

Automatically generated - may contain errors

Lab products found in correlation

Lipopolysaccharides (lps), by Merck Group (6 mentions) Interleukin 4 (il 4), by Thermo Fisher Scientific (5 mentions) Ionomycin, by Merck Group (4 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (4 mentions) Il 21, by Thermo Fisher Scientific (4 mentions) Human α thrombin, by Enzyme Research (3 mentions) E coli lps, by Merck Group (2 mentions) Ack lysing buffer, by Lonza (2 mentions) Rhil 21, by Thermo Fisher Scientific (2 mentions) Fortessa flow cytometer, by BD (2 mentions) Anti cd3 cd28 beads, by Thermo Fisher Scientific (2 mentions) Fetal bovine serum (fbs), by Gemini Bio (2 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions) Phorbol myristate acetate (pma), by Merck Group (2 mentions) P gingivalis lps, by InvivoGen (2 mentions) Cd40l, by Abcam (2 mentions) Il 21, by R&D Systems (2 mentions) Il 21, by Miltenyi Biotec (2 mentions) Human b cell isolation kit 2, by Miltenyi Biotec (2 mentions) Anti igm, by Jackson ImmunoResearch (2 mentions)

51 protocols using cd40l

B cell stimulation and IL-10 response

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

B cell number was counted by hemacytometer. Each 1×10⁶ B cells were cultured in 200 μL IMDM+GlutaMAX^TM (Life Technologies, Carlsbad, CA, USA) complete medium (contains 10% FCS, 100 U/mL penicillin, 100 mg/mL streptomycin, 2 mM L-glutamine, 2.5 μg/mL Amphotericin B and 50 μM 2-ME) in 96-well plates under the following conditions: control, CD40L, CD40L+LPS, CD40L+CpG, or CD40L+LPS+CpG in the absence or in the presence of fixed P. gingivalis. Final concentrations of these stimulants were as follows: CD40L (eBioscience, San Diego, CA, USA, 1 μg/mL), P. gingivalis LPS (Invivogen, San Diego, CA, USA, 10 μg/mL), mouse CpG-DNA (Hycult, Plymouth Meeting, PA, USA, 10 μM), and fixed P. gingivalis (5×10⁶/1×10⁶ cells). P. gingivalis LPS was used as TLR4 agonist and mouse CpG-DNA(5’-TCCATGACGTTCCTGATGCT -3’) was used as TLR9 agonist. B cells cultured without stimulation were used as control. Cells were cultured in a humidified incubator at 37°C with 5% CO₂ for 48 h. Thirty of 200 μL medium in each well was used to determine CD1d^highCD5⁺ B cell percentages and remaining cells were used to determine IL-10 mRNA expression levels. Culture supernatant was used for secreted IL-10 levels measurement.

LIU Z., HU Y., YU P., LIN M., HUANG G., KAWAI T., TAUBMAN M., WANG Z, & Xiaozhe H. (2017). Toll-like receptor agonists Porphyromonas gingivalis LPS and CpG differentially regulate IL-10 competency and frequencies of mouse B10 cells. Journal of Applied Oral Science, 25(1), 90-100.

+ Open protocol

+ Expand

Activation and Tracking of PBMCs

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

PBMCs from the Red Cross cohort were thawed and incubated with CFSE (Life Technologies, Carlsbad, CA) for 10 min as per manufacturer’s instructions. 5 mL of FBS (Gemini Bioproducts, West Sacramento, CA) was added and cells were incubated on ice for 5 min. Cells were rinsed and cultured with 10 μg/mL (Invivogen, San Diego, CA), 10 μg/mL CD40L (Invitrogen, San Diego, CA) and 2.5 ng/mL rhIL-21(Thermo Fisher Scientific), or CpG with CD40L and rhIL-21 similar to conditions described by Marasco et al. [11 (link)]. Controls with no stimuli, no CFSE, and anti-CD3/CD28 beads (Thermo Fisher Scientific) were included with each culture. Cells were then surface stained and acquired via flow cytometry using the Fortessa flow cytometer (BD Biosciences).

Nipper A.J., Smithey M.J., Shah R.C., Canaday D.H, & Landay A.L. (2018). Diminished antibody response to influenza vaccination is characterized by expansion of an age-associated B-cell population with low PAX5. Clinical immunology (Orlando, Fla.), 193, 80-87.

+ Open protocol

+ Expand

Induction of Enteric Glial Cell Sepsis

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

To induce the cell sepsis model, EGCs were stimulated by single lipopolysaccharide (LPS) (10 μg/mL, E. coli O111:B4, Sigma-Aldrich, St. Louis, MO, USA), CD40L (1 μg/mL, PeproTech, Rocky Hill, NJ, USA) or LPS + CD40L. After 24 h, the cell culture supernatants were collected as EGCs-conditioned medium for cytokine measurement and Caco-2 cell treatment.
For the siRNA transfection, CD40-siRNA, TRAF6-siRNA, negative control siRNA (NC-siRNA), and siRNA-Mate transfection reagent were purchased from GenePharma Co. Ltd, (Shanghai, China). EGCs were transfected with CD40-siRNA, TRAF6-siRNA or NC-siRNA using siRNA-Mate according to the manufacturer’s protocols for 72 h before LPS and CD40L stimulation.

Cheng B., Du M., He S., Yang L., Wang X., Gao H., Chang H., Gao W., Li Y., Wang Q, & Li Y. (2022). Inhibition of platelet activation suppresses reactive enteric glia and mitigates intestinal barrier dysfunction during sepsis. Molecular Medicine, 28, 137.

+ Open protocol

+ Expand

Induction of Enteric Glial Cell Sepsis

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

+ Open protocol

+ Expand

Activation of Naïve B Cells into Plasmablasts

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

For 24 or 48 h cultures, PBMC was thawed and stimulated with IL-21 (50 ng/mL) (Miltenyi) with and without CD40L (5 μg/mL) (Peprotech). Four hours prior to harvest, brefeldinA/monensin were added to the cultures. Cells were harvested and stained for flow cytometry.
For plasmablast cultures, naïve B cells were isolated using the human naïve B cell isolation kit II (Miltenyi) and purity was assessed following 12-day cultures. Samples with purity >90% were plated at two million cells per well and cultured with IL-21 (15 pg/mL or 50 ng/mL) (Miltenyi) and CD40L (5 μg/mL) (Peprotech).

Dam E.M., Maier A.C., Hocking A.M., Carlin J., Ng B, & Buckner J.H. (2018). Increased Binding of Specificity Protein 1 to the IL21R Promoter in B Cells Results in Enhanced B Cell Responses in Rheumatoid Arthritis. Frontiers in Immunology, 9, 1978.

+ Open protocol

+ Expand

CD40L Stimulation for B Cell Activation

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

All antibodies used are listed in Ting Information Table S1. Pacific Blue and Alexa 750 dyes used for barcoding (6 ) were from Molecular Probes. rhCD40L was from Peprotech (0.05 μg/ml) or Enzo Life Siences (0.25 μg/ml). CD40L from Enzo is fused to a FLAG® tag and was pre-incubated for 30 min with an Enhancer for ligands (anti-FLAG antibody; 1 μg/ml) to enhance the biological activity, according to manufacturer’s recommendations. CD40L from Enzo was used at a saturating concentration which potently induces proliferation and plasma cell differentiation in vitro when combined with IL-21 (Supporting Information Fig. S1). CD40L from Peprotech gave similar signaling responses without crosslinking (Supporting Information Fig. S2). CD40L from Enzo was used for Figure 2 and 4, CD40L from Peprotech was used for Figure 3.

Huse K., Wogsland C.E., Polikowsky H.G., Diggins K.E., Smeland E.B., Myklebust J.H, & Irish J.M. (2019). Human germinal center B cells differ from naïve and memory B cells in CD40 expression and CD40L-induced signaling response. Cytometry. Part A : the journal of the International Society for Analytical Cytology, 95(4), 442-449.

+ Open protocol

+ Expand

Splenocyte Activation Assay with CD40L and LPS

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

Mice were euthanized in a CO₂ chamber and single-cell suspensions of splenocytes were obtained by dispersing spleen tissues through a 60-gauge stainless steel screen. Erythrocytes were removed by ACK lysing buffer (Lonza, MA). Isolated splenocytes were added into 96-well plates (2.0 × 10⁵/well) in 200 µl RPMI complete medium containing 10% FBS. CD40L (Thermo Scientific) and E. coli LPS (strain O55:B5, Sigma-Aldrich) were used as agonist. Splenocytes from WT and TLR4^−/− mice were divided into 4 treatment groups: control; E. coli LPS (10 µg/ml); CD40L (1 µg/ml); E. coli LPS (10 µg/ml) + CD40L (1 µg/ml). Cells were cultured at 37°C in a humidified incubator with 5% CO₂ for 2 days and then were collected for analysis.

Lin M., Lin J., Wang Y., Bonheur N., Kawai T., Wang Z, & Han X. (2015). Lipopolysaccharide Attenuates CD40 Ligand-Induced Regulatory B10 Cell Expansion and IL-10 Production in Mouse Splenocytes. Open journal of immunology, 5(1), 1-8.

+ Open protocol

+ Expand

Flow Cytometry of CD40 Activated PBMCs

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

Fresh human PBMCs were cultured with different concentrations of MAB273, CP-870,893 or CD40L (ThermoFisher, 34-8902-81) for 20 min at 4 °C and were then washed with cold PBS prior to incubation with 1 μg/mL CD40L-biotin (ThermoFisher, 15836427) for 20 min at 4 °C. Cells were washed with cold PBS and LIVE/DEAD staining as well as cell surface staining with a panel of fluorescently labeled antibodies was performed as described above (Table S1). After staining and washing, PBMCs were resuspended in 1% PFA and acquired on an LSRFortessa flow cytometer.

Yan X., Ols S., Arcoverde Cerveira R., Lenart K., Hellgren F., Ye K., Cagigi A., Buggert M., Nimmerjahn F., Falkesgaard Højen J., Parera D., Pessara U., Fischer S, & Loré K. (2023). Cell targeting and immunostimulatory properties of a novel Fcγ-receptor-independent agonistic anti-CD40 antibody in rhesus macaques. Cellular and Molecular Life Sciences, 80(7), 189.

+ Open protocol

+ Expand

Lentiviral Transduction of Splenocytes

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

A total of 1 × 10⁶ splenocytes were stimulated with 5 µg/mL of LPS (InvivoGen) for 2 or 4 d in complete culture medium composed of Roswell Park Memory Institute medium (RPMI) supplemented with 10% fetal calf serum (Sigma), 0.05 mM 2-mercaptoethanol, 100 U/mL penicillin–streptomycin, 1 mM sodium pyruvate, and 0.1 mM nonessential amino acids (Gibco). Where indicated, cells were treated with 20 μM M1 and 10 μM Mdivi (Sigma) for 2 d from day 2 to day 4.
Stx5a-specific shRNAs were designed thanks to the RNAi consortium (https://portals.broadinstitute.org/gpp/public) and cloned in the pLKO.3G (Addgene #14748) vectors. Lentiviral particles were produced in the Human Embryonic Kidney 293T (HEK293T) cell line with the psPAX2 (Addgene #12260) and pMD2.G (Addgene #12259) vectors. For primary cell transduction, splenocytes were put in culture in complete culture medium in the presence of 80 ng/mL CD40L (Thermo Scientific) and 1 U/mL interleukin-4 (Miltenyi) for 24 h to promote B cell entry into cycle. The cells were then washed and transduced with lentiviral particles together with polybrene. After 24 h, cells were washed and differentiated into PCs by addition of LPS as indicated above.

Bonaud A., Gargowitsch L., Gilbert S.M., Rajan E., Canales-Herrerias P., Stockholm D., Rahman N.F., Collins M.O., Taskiran H., Hill D.L., Alloatti A., Alouche N., Balor S., Soldan V., Gillet D., Barbier J., Bachelerie F., Smith K.G., Jellusova J., Bruhns P., Amigorena S., Balabanian K., Linterman M.A., Peden A.A, & Espéli M. (2023). Sec22b is a critical and nonredundant regulator of plasma cell maintenance. Proceedings of the National Academy of Sciences of the United States of America, 120(2), e2213056120.

+ Open protocol

+ Expand

Autologous T-cell Response Assay to Tumor-Loaded DCs

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

Autologous mixed leukocyte reactions (MLRs) were performed to assay T-cell responses to DC that had been antigen-loaded by phagocytosis during co-culture with irradiated tumor cells (TC). Autologous PBMC were co-cultured with autologous TC or autologous DCV at PBMC:DCV and PBMC:TC ratios of 3:1 (6 million PBMC to 2 million antigen-loaded DC, or 6 million PBMC to 2 million TC) on a 6-well ultra-low attachment plate. The co-cultures were incubated at 37°C and 5% CO₂ for 6 days. On day-6, the PBMCs were re-challenged with another 2 million TC or 2 million antigen-loaded DC, respectively. The co-cultures were analyzed by flow cytometry on day 12. Control MLR groups included PBMC alone, non-antigen-loaded DC, PBMCs treated with phorbol-12-myristate-13-acetate (PMA, 50 ng/ml, Sigma-Aldrich, St. Louis, MO) and ionomycin (1 μg/ml, Sigma-Aldrich, St. Louis, MO), PBMCs co-cultured with antigen-loaded DC in the presence of costimulatory molecule CD40L (500 ng/ml, Thermo Fisher Scientific, Waltham, MA) and anti-human CD28/CD49d antibodies (500 ng/ml, BD Biosciences, San Jose, CA), and PBMCs co-cultured with antigen-loaded DC in the presence of anti-IL-12 antibody (LEAF, 100 μg/ml, BioLegend, San Diego, CA).

Dillman R.O., Nistor G.I, & Poole A.J. (2019). Genomic, proteomic, and immunologic associations with a durable complete remission of measurable metastatic melanoma induced by a patient-specific dendritic cell vaccine. Human Vaccines & Immunotherapeutics, 16(4), 742-755.

+ 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!