Immunoglobulin VH usage analysis by fluorescent in situ hybridization and flow cytometry
- PMID: 1517596
- DOI: 10.1016/0022-1759(92)90328-q
Immunoglobulin VH usage analysis by fluorescent in situ hybridization and flow cytometry
Abstract
We have devised a flow cytometry-based fluorescent in situ hybridization assay that permits analysis of gene expression in a large number of single cells. In this technique, fixed and permeabilized cells are incubated with biotinylated single-stranded RNA probes and by means of a fluorescently labelled second-step reagent, the cells are analyzed by flow cytometry. This is a rapid and simple method that allows all of the steps in the procedure to be performed on cells in suspension. Using this approach, we demonstrate here that immunoglobulin heavy chain variable region (VH) gene expression can be analyzed among individual cells using particular VH family-specific probes. This technique has a high degree of accuracy (greater than 97%) in detecting the fraction of cells expressing a specific message in a population and is sensitive enough to detect immunoglobulin message in LPS activated B cells. The technique has been applied successfully to monitor gene expression in homogeneous and heterogeneous populations. It also allows concurrent analysis of cell surface proteins and gene expression through two-color flow cytometry. This method of monitoring gene expression in individual cells may have a number of applications in immunology and cell biology.
Similar articles
-
Identification of malignant cells in multiple myeloma bone marrow with immunoglobulin VH gene probes by fluorescent in situ hybridization and flow cytometry.J Clin Invest. 1995 Mar;95(3):964-72. doi: 10.1172/JCI117805. J Clin Invest. 1995. PMID: 7883997 Free PMC article.
-
Strain-dependent expression of VH gene families.J Immunol. 1988 Apr 1;140(7):2436-41. J Immunol. 1988. PMID: 3127467
-
Stochastic pairing of heavy-chain and kappa light-chain variable gene families occurs in polyclonally activated B cells.Proc Natl Acad Sci U S A. 1990 Jul;87(13):4932-6. doi: 10.1073/pnas.87.13.4932. Proc Natl Acad Sci U S A. 1990. PMID: 2114644 Free PMC article.
-
Differential expression of VH gene families in peripheral B cell repertoires of newborn or adult immunoglobulin H chain congenic mice.J Exp Med. 1992 Jun 1;175(6):1449-56. doi: 10.1084/jem.175.6.1449. J Exp Med. 1992. PMID: 1588274 Free PMC article.
-
[Structure and function of the immunoglobulin Vh gene in man].Allerg Immunol (Leipz). 1990;36(4):225-32. Allerg Immunol (Leipz). 1990. PMID: 2129098 Review. German.
Cited by
-
Identification of malignant cells in multiple myeloma bone marrow with immunoglobulin VH gene probes by fluorescent in situ hybridization and flow cytometry.J Clin Invest. 1995 Mar;95(3):964-72. doi: 10.1172/JCI117805. J Clin Invest. 1995. PMID: 7883997 Free PMC article.
Publication types
MeSH terms
Substances
Grants and funding
LinkOut - more resources
Full Text Sources
Other Literature Sources