Related Experiment Video
Updated: Aug 10, 2026
Rapid and Robust Analysis of Cellular and Molecular Polarization Induced by Chemokine Signaling
Published on: December 12, 2014
Methods for phenotyping polarized and locomotor human lymphocytes
1Immunology Department, University of Glasgow (Western Infirmary), UK.
Insights
Researchers developed new methods to assess lymphocyte phenotype and locomotion simultaneously. These techniques are crucial for understanding mixed cell populations and immune responses.
Area of Science:
- Immunology
- Cell Biology
Background:
- Lymphocytes exhibit diverse phenotypes and motility.
- Studying lymphocyte locomotion requires methods that can analyze mixed cell populations, as cell interactions may influence movement.
Purpose of the Study:
- To develop and validate in vitro methods for simultaneously assessing lymphocyte phenotype and locomotor activity.
- To enable the study of lymphocyte behavior within complex cellular environments.
Main Methods:
- Adaptation of the lymphocyte polarization assay using alkaline phosphatase-anti-alkaline phosphatase (APAAP) staining after glutaraldehyde fixation.
- Development of a method where lymphocytes invade collagen gels, followed by collagenase digestion to isolate motile cells for immunofluorescence phenotyping via fluorescence-activated cell sorting (FACS).
Main Results:
- Successful adaptation of the lymphocyte polarization assay for simultaneous assessment.
- Collagenase digestion effectively recovered motile lymphocytes without altering the immunofluorescence staining patterns of common markers (CD3, -4, -8, -19, -29, -45RO, -45RA).
Conclusions:
- The described methods provide novel tools for analyzing lymphocyte phenotype and locomotion concurrently.
- These techniques are valuable for investigating lymphocyte behavior in mixed cell populations and understanding immune cell interactions.
Abstract:
Lymphocytes show heterogeneity both in phenotype and in locomotor activity; methods which permit the simultaneous assessment of both are therefore useful. The activation of locomotion may be dependent on interactions between lymphocytes and accessory cells, making it necessary to study mixed cell populations. We describe here two in vitro procedures which do this. Firstly the lymphocyte polarization assay has been adapted by using the alkaline phosphatase-anti-alkaline phosphatase method (APAAP) after fixation with glutaraldehyde. Secondly, we have allowed lymphocytes to invade collagen gels and then digested the gel with collagenase to recover the locomotor population. This can then be phenotyped by immunofluorescence using a fluorescence-activated cell sorter (FACS). Collagenase did not affect the staining pattern with commonly-used markers such as CD3, -4, -8, -19, -29, -45RO and -45RA.
