JoVE Journal

Immunology and Infection

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Application of Long-term cultured Interferon-γ Enzyme-linked Immunospot Assay for Assessing Effector and Memory T Cell Responses in Cattle

12.1K Views

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15:57 min

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July 11th, 2015

DOI :

10.3791/52833-v

July 11th, 2015

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Mayara F. Maggioli¹^,², Mitchell V. Palmer¹, H. Martin Vordermeier³, Adam O. Whelan³, James M. Fosse⁴, Brian J. Nonnecke¹, W. Ray Waters¹

¹Infectious Bacterial Diseases Research Unit, National Animal Disease Center, Agricultural Research Service, United States Department of Agriculture, ²Department of Veterinary Pathology, College of Veterinary Medicine, Iowa State University, ³UK Veterinary Laboratories Agency, ⁴Visual Services, National Centers for Animal Health, Animal and Plant Health Inspection Service, United States Department of Agriculture

Summary

Long-term cultured interferon-γ enzyme-linked immunospot assay is used as a measure of central memory responses and correlates with protective anti-mycobacterial vaccine responses. With this assay, peripheral blood mononuclear cells are stimulated with mycobacterial antigens and interleukin-2 for 14 days, enabling differentiation and expansion of central memory T cells.

Explore More Videos

Effector T Cells

Central Memory T Cells

Effector Memory T Cells

Antigen Stimulation

Long term Culture

Chapters in this video

0:00

Title

1:40

Methods

2:05

Long-term Culture

3:38

Elispot Assay Day 1

4:30

Elispot Assay Day 2

5:15

Adherent Cell Isolation

5:37

Flow Cytometry Plate Preparation

6:15

Harvest the 13 Day Long-term Cells

9:05

Elispot Assay Day 3

10:04

Alkaline Phosphatase Preparation

11:08

Substrate Solution Preparation

11:55

Reading the Plate

12:12

Performing Flow Cytometry

12:25

Results

14:37

Conclusions

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