Comparing And Contrasting Techniques That Measure Cell Mediated Immunity Discussion Forum

London, London,
Friday, 06 July 2012
Discussion Workshop:

Comparing and Contrasting Techniques that Measure Cell-mediated Immunity 

An event from European Scientific Conferences - Euroscicon "Specialising in communicating cutting edge technology & methodology in the Life Sciences"  Linked In

Comparing And Contrasting Techniques That Measure Cell Mediated Immunity Discussion Forum
Friday, 06 July 2012 09:00 - 17:00

The Royal College of Pathologists
Watson & Crick Room
2 Carlton House Terrace
United Kingdom

Map and Directions

This event is a discussion forum, focused on comparing and contrasting available techniques that measure cell mediated immunity. The study of cell mediated immunity requires the identification of cell types and subsets involved, accurate determination of levels of effector molecules, and the kinetic assessment of these same effector molecules in real time. Advances in Fluorescence Activated Cell Sorting (FACS) allow the assessment of multiple effector molecules, via intracellular antigen detection, in defined cell subsets, via extracellular antigen detection. However, it does not allow the precise and timely assessment of these same factors, especially after re-stimulation of cell samples. Enzyme-Linked ImmunoSorbent Assay ELISA and Cytometric Bead Array (CBA) allow the accurate determination of one or multiple effector molecules, but information on their cellular origin is often sacrificed. Participants will have a chance to explore the advantages and pitfalls of effector molecule detection with the experts during round table and panel discussions


Meeting Chair: Dr Marc Veldhoen, The Babraham Institute, Cambridge, UK


This event has CPD accreditation

On registration please submit your questions to the panel that will be asked by the chair on the day of the event


9:00 – 9:30          Registration

9:30 – 9:35          Introduction by Meeting Coordinator: Dr  Astrid Englezou, EuroSciCon, London, UK

9:35 – 9:45          Introduction by the Chair: Dr Marc Veldhoen, The Babraham Institute, Cambridge, UK


Talks by Invited Experts:


9:45 – 10:15        Flow cytometry and cytokine detection

Dr Marc Veldhoen

Babraham Institute, Cambridge, UK


10:15 – 10:45      Use of Multiplex Cytokine Analysis in Determining Secretion Profiles of Leucocyte Populations in the Female Reproductive Tract

Dr Gendie Lash

Lecturer at Newcastle University

Leucocytes are a major cell population in the non-pregnant endometrium and early pregnancy decidua.  Some of these cells have non-immune functions in addition to their classical immune functions.  To decipher the functions of two of these cell types, uterine macrophages and uterine natural killer (uNK) cells we have used multiplex cytokine and growth factor analysis to determine the secretion profiles of these cell types.  This has led to hypothesis driven functional studies based on their secretion profiles.  Data will be presented on initial studies into choice of multiplex analysis system and the type of data we are able to generate using this approach compared to more conventional ELISA.


10:45 – 11:15       Mid-morning Break


11:15 – 11:45      Fluorescently labeled tetrameric MHC-peptide complexes
Dr Stuart Sims, Nuffield Department of Medicine and NIHR Biomedical Research Centre University of Oxford Oxford UK

The development of the fluorescently labeled tetrameric MHC-peptide complex has enabled the direct visualisation, quantification and phenotypic characterisation of antigen-specific T cells using flow cytometry and has transformed our understanding of cellular immune responses. The combination of this technology with functional assays provides many new insights into these cells, allowing investigation into their lifecycle, manner of death and effector function. This talk will provide an overview of the techniques used in the construction of these tetramers, the problems and solutions associated with them, and the methods used in the study of antigen-specific T cells. Understanding how the antigen-specific cells develop and function in different circumstances and with different pathogens will be key to understanding natural host defence, as well as vaccine design and assessment.


11:45 – 12:15       The use of flow cytometry to dissect vaccine and pathogen induced T cells responses

Dr Phil Hogarth, TB Immunology, Animal Health & Veterinary Laboratory Agency¸UK

Using a murine model of BCG vaccination & M. bovis challenge, we are investigating the T cell responses responsible for providing (driving?) protective immunity. Traditional techniques such as ELISPOT & ELISA combined provide valuable data, but fail to identify the phenotype or multiple functionality of responder T cells. Using Intracellular Staining (ICS) techniques, we are able to generate data describing 7 separate subsets of T cells based on cytokine functionality combined with surface phenotype.

We demonstrate that a single systemic BCG vaccination induces distinct systemic and mucosal populations of T effector memory (TEM) cells in vaccinated mice. These CD4+CD44hiCD62LloCD27- T cells are maintained for long periods in BCG protected mice, maintaining a vaccine–specific functionality. 


12:15 – 12:45       FluoroSpot analysis of TLR-activated monocytes reveals several distinct cytokine secreting subpopulations

Christian Smedman, Mabtech AB, Sweden

A hallmark property of monocytes is their ability to rapidly produce and secrete a broad range of cytokines in response to TLR-stimulation. Earlier investigations of this diversity have for the most part been based on methods such as ELISA or Luminex, making it difficult to draw conclusions as to their functional profile on the cellular level. Mabtech has developed a range of FluoroSpot reagents which allows detection and quantification of cells secreting two cytokines simultaneously. By this technique it is possible to identify several subpopulations of monocytes with distinct cytokine secreting profiles in response to ligands against TLR2, TLR4 and TLR7/8. This talk will describe the FluoroSpot methodology which offers a sensitive and robust method for detecting dual cytokine secretion at the single cell level. Possible implications of the results to the general field of monocyte and macrophage biology will also be discussed


12: 45 – 13:15      Working Lunch

Please collect your lunch and take it to your discussion table

This is also a good time to fill out your feedback forms


13:15 – 14:45      Discussion Group Sessions

  • Round table discussion groups (20 minutes each) will be held throughout the afternoon
  • Delegates will rotate so that they may participate in all the discussion tables
  • All delegates will also be allocated a session for visiting the exhibition stands
  • Where appropriate delegates will be able to bring their samples to the discussions
  • See end of agenda for description of discussion tables


14:45 – 15:15      Afternoon Tea


15:15  – 16:15      Question and Answer Session

This session will include summing up of the discussion tables and questions submitted both prior to the meeting and throughout the day


16:15                     Chairman’s Summing Up and Feedback Prize Draw


About the Speakers

Phil Hogarth started his research career at the Liverpool School of Tropical Medicine, with a PhD in Immunoparasitology. Following postdoctoral training at Bristol University he joined AHVLA (formerly VLA) in 2001, becoming a team Leader/Senior Scientist. His main research interests are the T cell mechanisms by which vaccination protect against TB, and the identification of correlates of vaccine induced protection. Phil’s expertise lies in the use of flow cytometry to identify the phenotype and function of T cells induced by vaccination or infection with TB. He is reviewer for many journals & funding agencies, including the EU and Wellcome Trust and the author of 23 papers.


Stuart Sims is a Wellcome Trust funded research assistant now enrolled for a DPhil. He is studying the transcriptional regulation of memory inflation and CD161 expression.


Gendie Lash, who obtained her undergraduate and PhD degrees in Biochemistry from University of Otago, Dunedin, New Zealand in 1997.  She then did Post-Doc jobs in Obstetrics and Gynaecology, University of Nottingham and Department of Anatomy and Cell Biology, Queen’s University, Kingston, Ontario, Canada where she held a Canadian Hypertension Society Post-Doctoral Fellowship.  In 2002 she moved to Newcastle University where she has been ever since and is currently a lecturer.  Her current research focuses on blood vessel development in the non-pregnant uterus and maternal adaptations to pregnancy, with particular interest in the functional role of uterine leucocytes in these processes.


Keywords:  Flow Cytometry, ICS, ELISPOT, BCG, TB, BCG, uNKCytokine, ELISA, Multiplex. TLR. Toll, TB, TLR2, TLR4, FluoroSpot

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