Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Experiment Videos

A fluorescent cellular adhesion assay using insect cell produced human VCAM1

J K Stoltenborg1, P W Tsao, H J George

  • 1Department of Cardiovascular Diseases, Du Pont Merck Pharmaceutical Company, Wilmington, DE 19880-0400.

Journal of Immunological Methods
|September 30, 1994
PubMed
Summary

Researchers developed a new assay to measure T cell adhesion to VCAM1. This method uses recombinant VCAM1 (rVCAM1) and Jurkat T cells, offering a tool to study cell adhesion in inflammation and metastasis.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Evaluating the Renal Safety of Investigational New Drugs: Where Should We Be Going?

Clinical pharmacology and therapeutics·2017
Same author

Qualification of cardiac troponins for nonclinical use: a regulatory perspective.

Regulatory toxicology and pharmacology : RTP·2013
Same author

Synthesis and pharmacology of modified amidine isoxazoline glycoprotein IIb/IIIa receptor antagonists.

Bioorganic & medicinal chemistry letters·2001
Same author

Implementation of a continuous, enzyme-coupled fluorescence assay for high-throughput analysis of glutamate-producing enzymes.

Analytical biochemistry·2000
Same author

Tumor biology: use of tiled images in conjunction with measurements of cellular proliferation and death in response to drug treatments.

Clinical cancer research : an official journal of the American Association for Cancer Research·2000
Same author

Human immunodeficiency virus type 1 mutations selected in patients failing efavirenz combination therapy.

Antimicrobial agents and chemotherapy·2000

Area of Science:

  • Immunology
  • Cell Biology
  • Biochemistry

Background:

  • Activated endothelium and dendritic cells express VCAM1, an immunoglobulin superfamily adhesion molecule.
  • Mononuclear leukocytes express integrin VLA4, the counterreceptor for VCAM1.
  • VCAM1-VLA4 interactions regulate inflammation, cancer metastasis, and atherosclerosis.

Purpose of the Study:

  • To develop a fluorescent adhesion assay for measuring T cell adhesion to recombinant human VCAM1 (rVCAM1).
  • To establish a protocol for partial purification of non-denatured rVCAM1 from insect cell membranes.
  • To validate the assay using Jurkat T cells and specific antibodies.

Main Methods:

  • Expressed non-denatured rVCAM1 in a baculovirus expression vector system (BEVS).
  • Partially purified rVCAM1 from infected Sf9 insect cell membranes.

Related Experiment Videos

  • Developed a fluorescent adhesion assay using rVCAM1-coated 96-well plates and Jurkat T cells.
  • Utilized monoclonal antibodies against VCAM1, VLA4, and LFA1 for blocking studies.

    • Main Results:

    • Jurkat T cells specifically adhered to rVCAM1 membrane preparations.
    • Adhesion was blocked by antibodies against VCAM1 or VLA4, but not LFA1.
    • Both unstimulated and IL-2 stimulated Jurkat cells showed functional VLA4 binding.
    • The assay demonstrated specificity for the VCAM1-VLA4 interaction.

    Conclusions:

    • A functional fluorescent adhesion assay for VCAM1-VLA4 interactions was successfully developed.
    • The assay utilizes partially purified, non-denatured rVCAM1 expressed via BEVS.
    • This method provides a tool to study T cell adhesion and can be adapted for other receptor-ligand pairs.