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Combined immunomagnetic cell sorting and ELISPOT assay for the phenotypic characterization of specific antibody-forming cells.
- Source :
-
Journal of immunological methods [J Immunol Methods] 1997 Apr 25; Vol. 203 (2), pp. 193-8. - Publication Year :
- 1997
-
Abstract
- A combination of immunomagnetic cell sorting and ELISPOT techniques has been evaluated to permit enrichment and characterization of antibody-secreting cells (ASC). Cell suspensions containing putative ASC were first incubated with magnetic microbeads coated with antibodies specific for a given cell surface marker. After separation of bead-cell clusters and free cells, the resulting cell populations were examined for the presence of ASC by an ELISPOT assay. As a model system, the expression of selected cell differentiation markers by human circulating ASC has been evaluated after parenteral tetanus vaccination and during the course of a Leishmania infection. Prior treatment of blood MNC with beads coated with antibodies to CD38, HLA-DR or CD19 permitted the isolation of virtually all blood ASC. Further, prior immunomagnetic removal of T (CD2+) cells from blood MNC, followed by isolation of CD38+ cells facilitated the detection of Leishmania major-specific ASC in all six patients examined, whereas parasite-specific ASC among unfractionated blood mononuclear cells could only be detected in 3 out of these six patients. Simple and rapid, this approach provides not only accurate estimates of the frequency of ASC within a given B cell population or subpopulation, but can also efficiently enrich functional ASC from complex cell suspensions and thus should be particularly useful in situations where ASC are present at low frequencies.
- Subjects :
- Adult
Animals
B-Lymphocyte Subsets classification
Humans
Immunoglobulin G biosynthesis
Immunophenotyping
Leishmania major immunology
Leishmaniasis, Cutaneous immunology
Leukocytes, Mononuclear classification
Mice
Tetanus Toxoid immunology
Antibody Specificity
Antibody-Producing Cells classification
Enzyme-Linked Immunosorbent Assay methods
Immunomagnetic Separation methods
Subjects
Details
- Language :
- English
- ISSN :
- 0022-1759
- Volume :
- 203
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- Journal of immunological methods
- Publication Type :
- Academic Journal
- Accession number :
- 9149813
- Full Text :
- https://doi.org/10.1016/s0022-1759(97)00030-6