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Trfk5

Manufactured by BD
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The TRFK5 is a laboratory equipment designed for performing various analytical tasks. It functions as a versatile instrument for researchers and scientists. The core function of the TRFK5 is to facilitate precise and reliable measurements and analysis within a laboratory setting. Detailed specifications and intended use are not available for this product.

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Lab products found in correlation

Ebio13a, by Thermo Fisher Scientific (3 mentions) R3 34, by BD (2 mentions) Brefeldin a, by Merck Group (2 mentions) Ionomycine, by Merck Group (2 mentions) Foxp3 staining kit, by Thermo Fisher Scientific (2 mentions) Anti cd86, by BioLegend (1 mentions) Anti cd16 32 antibody, by BioLegend (1 mentions) Live dead fixable staining kits, by Thermo Fisher Scientific (1 mentions) Anti cd25, by BioLegend (1 mentions) Mp6 xt22, by BioLegend (1 mentions) Transcription factor staining buffer set, by Thermo Fisher Scientific (1 mentions) Monensin, by BioLegend (1 mentions) Bvd6 24g2, by BD (1 mentions) Bvd4 1d11, by BD (1 mentions) Tc11 18h10, by BD (1 mentions) Il 17a tc11 18h10, by BD (1 mentions) Phorbol myristate acetate (pma), by Merck Group (1 mentions)

6 protocols using trfk5

1

Inhibition of IL-5 and CTSL in Mice

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In mice, the inhibition of IL-5 was accomplished by administering an intravenous injection of an anti-IL-5 antibody (TRFK5; BD Pharmingen) or an isotype-matched control antibody (R3-34; BD Pharmingen) at a dose of 200 g/mouse, 24 h before PPE challenge. For CTSL inhibition, mice were pre-treated with 500 μg of CTSL inhibitor (SID 26681509, quarterhydrate) or a vehicle control, 12 h before PPE instillation. On the 3rd day after the final PPE challenge, the mice were euthanized, and samples were collected for subsequent analysis.
Xu X., Yu T., Dong L., Glauben R., Wu S., Huang R., Qumu S., Chang C., Guo J., Pan L., Yang T., Lin X., Huang K., Chen Z, & Wang C. (2023). Eosinophils promote pulmonary matrix destruction and emphysema via Cathepsin L. Signal Transduction and Targeted Therapy, 8, 390.
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2

Multiparametric Immune Cell Profiling

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Cells were incubated with anti-CD16/32 antibody (93; BioLegend) prior to antibody staining in order to prevent unspecific binding. LIVE/DEAD Fixable Staining Kits (Thermo Fisher Scientific) were used to exclude dead cells. For cell surface analysis, cells were stained with the following antibodies: anti-TCRβ (PE-Cy7/PE; H57–597), anti-KLRG1 (PE/BV605; 2F1/KLRG1), anti-CD25 (PE/PE-Cy7; PC-61), anti-CD86 (APC-Cy7; PO3.1), anti-MHCII (FITC; M5/114.15.2; all BioLegend) and anti-CD80 (PE; 16-10A1; ThermoFisher Scientific). For intracellular and intranuclear staining, cells were re-stimulated with phorbol myristate acetate (20 ng/ml) and ionomycin (1 µg/ml) for 6 hours with the addition of brefeldin A (1 µg/ml; all Sigma Aldrich) and monensin (2 µM; BioLegend) after 60 min. After surface and Live/Dead staining, cells were fixed using the Transcription Factor Staining Buffer Set (eBioscience) and incubated in Permeabilization buffer with antibodies specific to IL-2 (PE; JES6-5H4), IL-4 (PerCP-Cy5.5; 11B11), IL-6 (PE; MPS-20F3), TNFα (PE/FITC; MP6-XT22), IFNγ (APC; XMG1.2), GATA3 (FITC; 16E10A23; all BioLegend), IL-5 (PE; TRFK5; BD Pharmingen), and IL-13 (Alexa Flour 488; eBio13A; ThermoFisher Scientific). Data were acquired using a BD LSRFortessa II (BD 172 Biosciences) and analyzed by FlowJo software (Tree Star, Ashland, OR, USA).
Steinmann S., Schoedsack M., Heinrich F., Breda P.C., Ochel A., Tiegs G, & Neumann K. (2020). Hepatic ILC2 activity is regulated by liver inflammation-induced cytokines and effector CD4+ T cells. Scientific Reports, 10, 1071.
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3

Maternal IL-5 Neutralization during Pregnancy

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ACLP+/− pregnant females were injected intravenously with a neutralizing antibody against IL-5, (TRFK5; BD Pharmingen) or an isotype control (R3-34; BD Pharmingen) for five consecutive days, using a dose of 100 μγ on E13.5-E14.5 and 150 μγ on E15.5-E17.5. The amount of antibody was calculated to achieve a fetal dose of 10mg/kg (the same dose that is typically used in an adult mouse (13 (link))) and an estimated maternal-fetal transmission of 2-3% (14 (link)). Fetal intestines and liver were analyzed on E18.5. In the case of fetal intervention, fetuses were injected with the same antibodies on E13.5 using a fetal intrahepatic injection as previously described (15 -17 (link)). Briefly, mothers were anesthetized and the uterus was exposed by performing a laparotomy. The fetuses were injected individually, through the translucent intact uterus, with the neutralizing antibody; controls were injected with the isotype control antibody or PBS in a volume of 5 μl, using pulled glass micropipettes. The laparotomy was closed in layers and the fetuses were analyzed on E18.5.
Frascoli M., Jeanty C., Fleck S., Moradi P.W., Keating S., Mattis A.N., Tang Q, & MacKenzie T.C. (2016). Heightened immune activation in fetuses with gastroschisis may be blocked by targeting IL-5. Journal of immunology (Baltimore, Md. : 1950), 196(12), 4957-4966.
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4

Cytokine and Chemokine Measurement by ELISA

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Culture supernatant cytokines (mouse IL-4, IL-5 and IFN-γ) were measured by two-site sandwich ELISA. The Abs for coating the plates and the biotinylated secondary mAbs were as follows: for IL-4, rat mAb anti-mouse IL-4 (BVD4-1D11, BD Biosciences, San Diego, CA) and biotinylated mAb anti-mouse IL-4 (BVD6-24G2, BD Biosciences); for IL-5, rat mAb anti-mouse/human IL-5 (TRFK5, BD Biosciences) and biotinylated mAb anti-mouse IL-5 (TRFK4, BD Biosciences); for IFN-γ, polyclonal rabbit anti-mouse IFN-γ Ab (prepared in our laboratory) and biotinylated rat mAb anti-mouse IFN-γ (XMG1.2, BD Pharmingen). HRPO-conjugated Streptavidin was purchased from Zymed (San Francisco, CA). Levels of human TNF-α were determined by an ELISA system that was developed in our laboratory. Levels of human IL-10 were determined with human IL-10 ELISA Ready-SET-Go kit (Thermo Fisher Scientific, Waltham, MA). Eotaxin and thymus and activation-regulated chemokine (TARC) were measured with human CCL11/Eotaxin DuoSet ELISA Development Systems (R&D Systems, DY320) and human CCL17/TARC DuoSet ELISA Development Systems (R&D Systems, DY364), respectively.
Kohno K., Koya-Miyata S., Harashima A., Ariyasu T, & Ushio S. (2018). NK-4 exerts selective regulatory effects on the activation and function of allergy-related Th2 cells. PLoS ONE, 13(6), e0199666.
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5

Intracellular Cytokine Staining for ILCs

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For intracellular cytokines staining of ILCs, single-cell suspensions from lung were incubated at 37°C with Brefeldin A in association or not with PMA (50 ng/ml, Sigma)/Ionomycine (500 ng/ml, Sigma) or 50 ng/ml of IL-12 and IL-18 for 4 hours before being surface stained, fixed and permeabilized (Foxp3 staining kit, eBiosciences). mAbs for cytokines staining included IFN-γ (XMG1.2, Biolegend), IL-17A (TC11-18H10, BD Biosciences), IL-5 (TRFK5, BD Biosciences), and IL-13 (eBio13A, eBiosciences). To block glycolysis during ex vivo stimulation, cells were incubated in the presence of 10mM 2-DG (Sigma).
Corral D., Charton A., Krauss M.Z., Blanquart E., Levillain F., Lefrançais E., Sneperger T., Vahlas Z., Girard J.P., Eberl G., Poquet Y., Guéry J.C., Argüello R.J., Belkaid Y., Mayer-Barber K.D., Hepworth M.R., Neyrolles O, & Hudrisier D. (2022). ILC precursors differentiate into metabolically distinct ILC1-like cells during Mycobacterium tuberculosis infection. Cell Reports, 39(3), 110715.
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6

Intracellular Cytokine Staining of ILCs

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For intracellular cytokines staining of ILCs, single-cell suspensions from lung were incubated at 37°C with Brefeldin A in association or not with PMA (50 ng/ml, Sigma)/Ionomycine (500 ng/ml, Sigma) or 50 ng/ml of IL-12 and IL-18 for 4 hours before being surface stained, fixed and permeabilized (Foxp3 staining kit, eBiosciences). mAbs for cytokines staining included IFN-g (XMG1.2, Biolegend), IL-17A (TC11-18H10, BD Biosciences), IL-5 (TRFK5, BD Biosciences), and IL-13 (eBio13A, eBiosciences) For HIF1a staining of ILCs, single-cell suspensions from lung were incubated at 37°C with DMOG (500µM) for 3 hours before being surface stained, fixed and permeabilized (Foxp3 staining kit, eBiosciences). In order to block glycolysis during ex vivo stimulation, cells were incubated in the presence of 10mM 2-DG (Sigma). Mtb was inactivated by incubation in PFA 4% for 2 hours at room temperature.
Corral D., Charton A., Krauss M.Z., Blanquart E., Levillain F., Lefrançais E., Sneperger T., Girard J., Eberl G., Poquet Y., Guéry J., Argüello R.J., Hepworth M.R., Neyrolles O., & Hudrisier D. (2021). Metabolic regulation of ILC2 differentiation into ILC1-like cells during Mycobacterium tuberculosis infection.
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