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Updated: Feb 17, 2026
Static Adhesion Assay for the Study of Integrin Activation in T Lymphocytes
Published on: June 13, 2014
Protocol for adhesion and immunostaining of lymphocytes and other non-adherent cells in culture
Matthew Tsang1, Jennifer Gantchev2, Feras M Ghazawi1
1Division of Dermatology, University of Ottawa, Ottawa, Ontario, Canada.
Insights
This study introduces a simple gravity sedimentation method for adhering lymphocytes in multi-well plates for immunofluorescent staining. This technique avoids specialized equipment and lengthy preparation, enabling efficient cell analysis.
Area of Science:
- Cell Biology
- Immunology
- Biotechnology
Background:
- Immunostaining of non-adherent cells typically requires cell adhesion to slides via cytocentrifugation or charged coatings.
- These standard methods necessitate specialized equipment or extensive preparation time, posing limitations for researchers.
Purpose of the Study:
- To develop a simplified, accessible method for immunofluorescent staining of lymphocytes within multi-well culture plates.
- To provide an alternative to existing techniques that require specialized equipment or significant preparation.
Main Methods:
- Described a novel technique for adhering lymphocytes using gravity sedimentation in phosphate-buffered saline (PBS) within multi-well plates.
- Cells were allowed to settle onto plastic well bottoms or glass coverslips for 30 minutes without specialized coatings or equipment.
Main Results:
- Gravity sedimentation in PBS, but not serum-containing medium, effectively adhered lymphocytes to surfaces within 30 minutes.
- The method achieved dense cell coverage (1 × 10^6 cells) and preserved cell morphology.
- Successfully applied the technique for immunofluorescent staining of primary human lymphocytes and immortalized lymphoma cells.
Conclusions:
- A straightforward, cost-effective method for preparing lymphocytes for immunofluorescent staining in multi-well plates has been established.
- This gravity sedimentation technique offers a practical alternative for researchers, simplifying cell preparation and analysis.
- The method is versatile, applicable to various lymphocyte types while maintaining cellular integrity.
Abstract:
Immunostaining of non-adherent cells is commonly performed after adhesion of cells onto microscope slides either using cytocentrifugation or with the help of charged coating substrates. These techniques, however, require either specialized equipment or significant preparation time. Here, we describe a method for immunofluorescent staining of lymphocytes within multi-well culture plates, where cells suspended in phosphate buffered saline (PBS) are adhered to either the plastic well bottom or glass coverslips by gravity sedimentation. This technique requires only common laboratory materials, no coating steps, and allows for densely adherent cell coverage with 1 × 106 cells. Our data show that suspension of cells in PBS, but not serum-containing growth medium, allows for adhesion to plastic or glass after 30 min of gravity sedimentation. We show that this method is applicable for immunofluorescent staining of both primary human lymphocytes and immortalized lymphoma cells, and that it preserves cell morphology.
