Identification of B and T cells in human spleen sections by infrared microspectroscopic imaging
Article first published online: 23 FEB 2005
Copyright © 2005 Wiley-Liss, Inc.
Cytometry Part A
Volume 64A, Issue 2, pages 53–61, April 2005
How to Cite
Krafft, C., Salzer, R., Soff, G. and Meyer-Hermann, M. (2005), Identification of B and T cells in human spleen sections by infrared microspectroscopic imaging. Cytometry, 64A: 53–61. doi: 10.1002/cyto.a.20117
- Issue published online: 17 MAR 2005
- Article first published online: 23 FEB 2005
- Manuscript Accepted: 16 DEC 2004
- Manuscript Revised: 27 AUG 2004
- Manuscript Received: 5 MAY 2004
- Marie Curie Intra-European Fellowship, Sixth EU Framework Program
- Volkswagen Foundation
- secondary lymphoid organs;
- lymphocyte identification;
- infrared spectroscopic imaging
Infrared spectroscopy probes the chemical composition and molecular structure of complex systems such as tissue and cells. Infrared spectroscopic imaging combines this spectral information with lateral resolution near the single-cell level. We analyzed whether this method is competitive with classic immunohistochemical methods for immunologic tissue and cells.
We recorded infrared microspectroscopic mapping datasets with a 90- × 90-μm2 aperture from a 3- × 3-mm2 unstained tissue area of human spleen. A secondary follicle containing a germinal center and a T zone were studied in more detail by infrared microspectroscopic imaging with lateral resolution near 5 μm. The results were compared with consecutive sections stained by immunoglobulin D antibodies. T and B lymphocytes were extracted from human blood and served as independent test samples.
Cluster analysis of infrared datasets produced images that distinguished anatomical features such as primary and secondary follicles, T zones, arteries, and spleen red pulp. The assignments could be confirmed in consecutive sections by immunohistochemical staining. Main spectral variances between T and B lymphocytes in high-resolution measurements were attributed to specific spectral contributions of DNA and cytosol.
Sensitivity and specificity of the infrared based methods are comparable to those of standard staining procedures for identification of B and T cells. However, infrared spectroscopic imaging can offer advantages in velocity, data throughput, and standardization because of minimal sample preparation. The results emphasize the potential of infrared spectroscopy as an innovative tool for the distinction of cell types, in particular in immunologic tissue. © 2005 Wiley-Liss, Inc.