Detecting Human IL-17+ T Cells

The ex vivo detection of human IL-17⁺ T cells was performed as previously described (de Beaucoudrey et al., 2008 (link)). In brief, PBMCs were purified on Ficoll-Paque PLUS (GE Healthcare) and resuspended in RPMI + 10% FBS (Invitrogen). We took aliquots of 10⁶ nonadherent cells/ml and either left them unstimulated or stimulated them with 40 ng/ml PMA (Sigma-Aldrich) + 10⁻⁵ M ionomycin (Sigma-Aldrich) for 12 h. All cells were treated with 1 µl/ml GolgiPlug for the final 12 h of culture. Surface staining was performed with PE-Cy5–conjugated anti–human CD3 (PE Biosciences) Ab, and intracellular staining was performed with Alexa Fluor 488–conjugated anti–human IL-17 (eBioscience) and PE-conjugated anti–human IFN-γ (BD) or PE-conjugated anti–human IL-22 (R&D Systems) Abs. Cells were analyzed with a FACScan machine and CellQuest software (both from BD).

Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link: Access Free Full Text.

Kreins A.Y., Ciancanelli M.J., Okada S., Kong X.F., Ramírez-Alejo N., Kilic S.S., El Baghdadi J., Nonoyama S., Mahdaviani S.A., Ailal F., Bousfiha A., Mansouri D., Nievas E., Ma C.S., Rao G., Bernasconi A., Sun Kuehn H., Niemela J., Stoddard J., Deveau P., Cobat A., El Azbaoui S., Sabri A., Lim C.K., Sundin M., Avery D.T., Halwani R., Grant A.V., Boisson B., Bogunovic D., Itan Y., Moncada-Velez M., Martinez-Barricarte R., Migaud M., Deswarte C., Alsina L., Kotlarz D., Klein C., Muller-Fleckenstein I., Fleckenstein B., Cormier-Daire V., Rose-John S., Picard C., Hammarstrom L., Puel A., Al-Muhsen S., Abel L., Chaussabel D., Rosenzweig S.D., Minegishi Y., Tangye S.G., Bustamante J., Casanova J.L, & Boisson-Dupuis S. (2015). Human TYK2 deficiency: Mycobacterial and viral infections without hyper-IgE syndrome. The Journal of Experimental Medicine, 212(10), 1641-1662.

Publication 2015

Ficoll-Paque PLUS PMA ionomycin PE-conjugated anti–human IFN-γ FACScan machine CellQuest software

Alexa fluor 488 Anti cd3 Cells Ficoll Human Ifn γ Il 17 Il 22 Intracellular Ionomycin T cells

Corresponding Organization : Université Paris Cité

Other organizations : Cornell University, Bursa Uludağ Üni̇versi̇tesi̇, Hôpital Militaire Moulay Ismail, National Defense Medical College, Masih Daneshvari Hospital, Shahid Beheshti University of Medical Sciences, University of Hassan II Casablanca, University of Mendoza, Garvan Institute of Medical Research, St Vincent's Clinic, National Institutes of Health, National Institute of Allergy and Infectious Diseases, Université Ibn-Tofail, Karolinska Institutet, Singapore General Hospital, Karolinska University Hospital, King Saud University, Universidad de Antioquia, Hospital Sant Joan de Déu Barcelona, Baylor University, Ludwig-Maximilians-Universität München, Friedrich-Alexander-Universität Erlangen-Nürnberg, Délégation Paris 5, Sorbonne Paris Cité, Kiel University, Hôpital Necker-Enfants Malades, Assistance Publique – Hôpitaux de Paris, Sidra Medical and Research Center, Tokyo Medical and Dental University, Howard Hughes Medical Institute

Top 5 similar protocols

Variable analysis

independent variables

Stimulation with 40 ng/ml PMA + 10^-5 M ionomycin for 12 h

dependent variables

Detection of human IL-17+ T cells

control variables

PBMC culture conditions (RPMI + 10% FBS)
Cell treatment with 1 µl/ml GolgiPlug for the final 12 h of culture

controls

Unstimulated PBMCs (negative control)

Annotations

Based on most similar protocols

Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.

As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.

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!