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Methods for diagnosis of immune responses against viruses

A virus and influenza virus technology, which is applied in the fields of medicine, virology and immunology, can solve the problems of the T cell response method of the influenza virus, the lack of intracellular processing of the peptide, and the detection of the T cell response of the inactivated virus.

Inactive Publication Date: 2015-03-11
由卫生福利和体育大臣代表的荷兰王国
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Problems solved by technology

In addition, exogenously added peptides lack intracellular processing of peptides prior to presentation by MHC molecules, thereby potentially missing responses that depend on intracellular post-translational modifications
Furthermore, natural processing allows presentation of peptides at native levels, which lead to physiologically relevant immune responses in vitro, whereas exogenously added peptides may lead to stimulation of physiologically irrelevant T cell responses
[0009] However, the prior art does not disclose methods for using inactivated viruses to detect T cell responses against viruses where the T cell responses reliably reflect responses of sufficient magnitude to be obtained with live viruses
The prior art in particular does not disclose such a method for detecting T cell responses against influenza virus

Method used

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  • Methods for diagnosis of immune responses against viruses
  • Methods for diagnosis of immune responses against viruses
  • Methods for diagnosis of immune responses against viruses

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Embodiment 1

[0051] 1.1. Materials and methods

[0052] 1.1.1 Inclusion of donors and isolation of PBMCs

[0053] Blood buffy coats from healthy individuals were obtained from the Sanquin Blood Bank North West Region according to guidelines for human experimentation (Project No. S03.0015-X). Additionally, peripheral blood mononuclear samples were obtained from 2 previously healthy individuals (51-year-old female, 55-year-old male) with laboratory-confirmed influenza A(H1N1)v infection, 13 and 19 days after symptom onset, respectively. cells (PBMC). All participants provided written informed consent before the study began. This study was approved by the Medical Ethics Committee of the University Medical Center Utrecht. Human PBMC were isolated by density centrifugation and cryopreserved at -130°C in 90% FCS (Hyclone, Logan, Utah) / 10% DMSO (Sigma-Aldrich St. Louis, USA) until analysis.

[0054] 1.1.2 Influenza virus

[0055] Dr. S. van der Werf (Institute Pasteur, Paris, France) generou...

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Abstract

The present invention relates to methods for diagnosis of a cellular immune responses against a virus using an inactivated virus. In the method of the invention a cellular immune response against the virus is detected in a subject by incubating PBMCs from the subject with a preparation of inactivated virus and subsequently detecting the expression of at least one T cell specific cytokine in the subject's PBMCs, preferably by flow cytometry. Advantageously in the method, inactivated virus is used for incubation with PBMCs from the subject so as to make feasible that the method is performed in laboratories without BSL-3 classification. Preferably the method is a method for detecting a CD4+ and / or CD8+ T cell response against an influenza virus by detecting expression of one or more of CD107, IFN-γ, IL-2, IL-10 and TNF-α, using formalin inactivated influenza virus. The invention further pertains to kits comprising components that are useful for detecting a cellular immune responses in a subject against a virus.

Description

field of invention [0001] The present invention relates to the field of medicine. The invention relates in particular to the fields of virology and immunology, more particularly the invention relates to methods for diagnosing cellular immune responses against viruses using inactivated viruses, such as influenza viruses. Background of the invention [0002] Research on highly pathogenic and pandemic influenza viruses requires a BSL-3 high containment facility. This facility limitation substantially affects laboratory potential. Therefore, there is a need for an assay capable of detecting immune responses against highly pathogenic influenza viruses that can be performed without the need for a BSL-3 high containment facility. [0003] By inactivating highly pathogenic and pandemic influenza viruses, these viruses can be used in laboratories that do not have BSL-3 classification. Currently, formalin chemical treatment is used to inactivate influenza virus during the productio...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/569
CPCG01N2333/525G01N2333/555G01N33/505G01N2333/54G01N33/56972G01N2333/70596G01N33/569G01N33/68
Inventor E·C·瑟图特W·M·刘
Owner 由卫生福利和体育大臣代表的荷兰王国
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