Abstract
Our understanding of the level and distribution of gene and protein expression in cells is a key component of modern cell biological and medical research. Detecting intracellular proteins with labeled antibodies or genes with labeled oligonucleotide sequences by fluorescence microscopy requires fixation of the target molecule in its natural distribution and the penetration of the probe to the target. This typically involves chemical fixation followed by a detergent treatment that renders the cell membrane permeable to a labeling antibody. The advantages of using quantum dots (QDs) over organic dyes to detect expression, such as high brightness, stability, and simplified multiple target labeling has been described in previous publications. However, QDs are structurally larger than organic dye probes and require different fixation and permeabilization conditions for optimum labeling. In the chapter, we describe several protocols for labeling proteins in nuclear, cytoplasmic, and membranous compartments with QD conjugates.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Larsson, L. I. (1988) Immunocytochemistry: Theory and Practice. CRC Press, Boca Raton, FL.
Jacobberger, J. W. (1991) Intracellular antigen staining: quantitative immunofluorescence. Methods 2, 207–218.
Nakamura, F. (2001) Biochemical, electron microscopic and immunohistological observations of cationic detergent-extracted cells: detection and improved preservation of microextensions and ultramicroextensions. BMC Cell Biol. 2, 10.
Callow, M. G., Zozulya, S., Gishizky, M. L., Jallal, B., and Smeal, T. (2005) PAK4 mediates morphological changes through the regulation of GEF-H1. J. Cell Science 118, 1861–1872.
Ness, J. M., Akhtar, R. S., Latham, C. B., and Roth, K. A. (2003) Combined tyramide signal amplification and quantum dots for sensitive and photostable immunofluorescence detection. J. Histochem. Cytochem. 51, 981–987.
Kozubeka, S., Lukáováa, E., Amrichováa, J., Kozubekb, M., Likováa, A., and Lotováa, J. (2000) Influence of cell fixation on chromatin topography. Anal. Biochem. 282, 29–38.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2007 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Ornberg, R.L., Liu, H. (2007). Immunofluorescent Labeling of Proteins in Cultured Cells With Quantum Dot Secondary Antibody Conjugates. In: Bruchez, M.P., Hotz, C.Z. (eds) Quantum Dots. Methods in Molecular Biology, vol 374. Humana Press. https://doi.org/10.1385/1-59745-369-2:3
Download citation
DOI: https://doi.org/10.1385/1-59745-369-2:3
Publisher Name: Humana Press
Print ISBN: 978-1-58829-562-0
Online ISBN: 978-1-59745-369-1
eBook Packages: Springer Protocols