Immunology

Research Challenges

The TBRU seeks to understand how the host immune system works to control the growth of M. tuberculosis and to identify immune components as markers of protective immunity during different stages of M. tuberculosis infection.

The primary objective of the Immunology Component of the TBRU is to conduct a coordinated, multi-disciplinary investigation of host immunological response mechanisms to M. tuberculosis in order to identify surrogate markers for clinical use.

Sub-objectives Include:

• Use various models to evaluate disease progression or protection in humans.
• Develop or test new in vitro assays for determining a person\x92s ability to prevent or control Tb infection.
• Identify host immune factors associated with latent infection and reactivation.
• Identify host immune factors associated with disease protection.

Laboratory Sites

Case Western Reserve University, Cleveland, OHIO
Joint Clinical Research Centre (JCRC) - Kampala, Uganda
South Africa Tuberculosis Vaccine Initiative Laboratory (SATVI)\x96 Cape Town, South Africa

Organization and Operation

Immunologic studies are initiated primarily at CWRU under the direction of Dr. Henry Boom. Pilot studies are also accomplished at the sites of clinical trials with technology transfer to improve the services of the immunology laboratories at these sites.

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JCRC Immunology Laboratory-Uganda

The immunology laboratory at the Joint Clinical Research Centre is a full service laboratory headed by Mr. Pierre Peters. This laboratory works to provide immunologic laboratory services and participates in studies to evaluate the field and laboratory performance of immunologic tests needed to support TBRU clinical and basic research programs in Uganda. The laboratory also assists the TBRU Specimen Repository in processing, storage and shipment of specimens collected during TBRU clinical trials and other studies. Specimens may be shipped to the US and other international sites to facilitate testing of samples with assays not yet available in Uganda.

Assays

The laboratory has a Miltenyi MACSQuant Flow Cytometer.  This instrument allows for 9 parameter, 7 color flow cytometry.  Multicolor phenotypic and intracellular analysis can easily be accomplished using this three laser instrument.

The Elisaspot assay is well established in the laboratory. We do not have the capability to read the resultant plates but we have had very good success at shipping the plates back to the US to be read on instruments here at CWRU and in the laboratories of our collaborators in Portland, Oregon.

Recently we have obtained a Life Technologies  Magpix multiplex system onsite.  With the many multiplex panels available we can take advantage of this technology to quickly and efficiently analyze the various specimens we have collected from our studies.

The mainstay of this laboratory is the ElISA Immuno Assay. We have tested thousands of samples in our laboratory using both commercial ELISA’s and matched pair antibody self derived assays. Although primarily we are looking at IFNγ as a marker of activation, we have the ability to assay for other cytokines, such as TNFα, as well.

Our laboratory staff has been trained through visits to other laboratories outside of Uganda as well as through visits by TBRU staff from the US and other investigators from around the world. They are a knowledgeable group of professionals who can deliver high level results with minimal infrastructure. All laboratory staff have visited the Cleveland TBRU laboratory for training.

Staff

The Director of the TBRU, Dr. Henry Boom leads the Immunology Component. Mr. Keith Chervenak is the direct link for the Cleveland lab to the Kampala lab. He visits Uganda several times a year to manage the laboratory aspects of the many studies being conducted through the immunology laboratory. In addition, he frequently introduces new technologies and assays as well as to provide a one on one opportunity for discussions about the various on going projects.

SATVI Laboratory-South Africa

The South Africa Tuberculosis Vaccine Initiative laboratory comprises an academic research component and a clinical trial trials component which both comply with Good Laboratory Practice. Further, the clinical trials component of the laboratory is ISO15189 compliant as determined by the South African National accreditation System, SE Dept Trade and Industries.

The laboratories consist of an approximately 150SqM reference immunology research laboratory based at the Institute of Infectious Diseases and Molecular Medicine in Cape Town and a 103SqM satellite laboratory site at the SATVI Clinical Trials Site in Worcester. The facilities comprise P2 immunology laboratories and molecular products processing areas, a Flow Cytometry area and a state of the art Cryo-facility. The Cryofacility has 8-80°C freezers and 13 Liquid Nitrogen tanks and currently stores more than 600,000 samples from various tuberculosis study-related cohorts. This is an invaluable resource to the scientific community. The integrity of stored samples is under continuous monitoring and is subject to strict quality control procedures. The SATVI Quality Management System is under continuous internal review and has performed well under external audit.

Assays

Laboratory assay tests range from standardized procedures such as peripheral blood mononuclear PBMC isolation and cryopreservation to novel whole blood intracellular cytokine assays.

Test procedures included in ISO15189 compliance are PBMC isolation and Cryopreservation, Whole Blood Intracellular Cytokine Assays (both short term and long term), Elispot and Quanitiferon®.

Other assays routinely performed in the lab are Multiparameter Flow Cytometry, Dilution dye (CFSE) cell proliferation assays, Quantitative Real Time PCR and Western blots.

Staff

The SATVI laboratory functions under the Directorship of Prof. Willem Hanekom with E. Jane Hughes as the Laboratory Project Manager and Marwou de Kock as the Worcester Laboratory Manager. Currently, SATVI has a staff complement of 30; consisting of a Management team, Post doctoral fellows, Scientists, Medical Technologists, a Laboratory Assistant and a number of PhD, Masters and Honours students. Laboratory staff activities are supported by a dedicated Administrative, Regulatory and Clinical team.

Staff at all TBRU sites are trained in Good Clinical Practice and Good Laboratory Practice.